patterns of resource allocation in caribbean coral reef sponges
TRANSCRIPT
PATTERNS OF RESOURCE ALLOCATION IN CARIBBEAN CORAL REEF
SPONGES
Wai Leong
A Thesis Submitted to the
University of North Carolina Wilmington in Partial Fulfillment
of the Requirements for the Degree of
Master of Science
Department of Biology and Marine Biology
University of North Carolina Wilmington
2009
Approved by
Advisory Committee
___Richard M. Dillaman__ ____Stuart R. Borrett_____
____Joseph R. Pawlik____
Chair
Accepted by
______________________
Dean, Graduate School
ii
TABLE OF CONTENTS
ABSTRACT……………………………………………………………………………. ..iv
ACKNOWLEDGEMENTS………………………………………………………………vii
LIST OF TABLES……………………………………………………………………… .ix
LIST OF FIGURES……………………………………………………………………. .. .x
CHAPTER 1: FRAGMENTS VERSUS PROPAGULES: REPRODUCTIVE TRADE-
OFFS FOR TWO CALLYSPONGIA SPP. FROM FLORIDA CORAL REEFS.................1
ABSTRACT......................................................................................................................2
INTRODUCTION ............................................................................................................3
MATERIALS AND METHODS......................................................................................5
RESULTS .........................................................................................................................8
DISCUSSION...................................................................................................................9
LITERATURE CITED ...................................................................................................12
CHAPTER 2: IS THERE A TRADE-OFF BETWEEN GROWTH AND DEFENSE
AMONG CARIBBEAN CORAL REEF SPONGES? ......................................................18
ABSTRACT....................................................................................................................19
INTRODUCTION ..........................................................................................................20
MATERIALS AND METHODS....................................................................................22
RESULTS .......................................................................................................................23
DISCUSSION.................................................................................................................25
LITERATURE CITED ...................................................................................................30
CHAPTER 3: PATTERNS OF RESOURCE ALLOCATION IN CARIBBEAN
SPONGES: IS THERE A TRADE-OFF BETWEEN REPRODUCTION AND
DEFENSE? ........................................................................................................................39
ABSTRACT....................................................................................................................40
iii
INTRODUCTION ..........................................................................................................41
MATERIALS AND METHODS....................................................................................44
RESULTS .......................................................................................................................45
DISCUSSION.................................................................................................................46
LITERATURE CITED ...................................................................................................52
iv
ABSTRACT
Trade-offs are a common theme in the ecological literature. Organisms allocate
resources to physiological functions such as growth and reproduction. When resources
are limiting, organisms must selectively allocate their resources, leading to resource
allocation trade-offs. Among Caribbean reef sponges, some species produce secondary
metabolites that deter predation. Yet, other species that do not produce any chemical
defenses co-exist alongside their defended counterparts. Resource allocation trade-offs
have been demonstrated to explain the co-existence of undefended and defended species
in terrestrial plants. Species either deter predation by allocating resources to defense, or
tolerate predation by allocating resources to growth and reproduction.
Although previous work has provided some evidence for resource allocation trade-
offs between chemical defense, growth and reproduction in Caribbean coral reef sponges,
this is the first work measuring growth rates and reproductive output in sponge species
for the purpose of comparing between the undefended and defended species most
commonly found on Caribbean coral reefs. First, a resource allocation trade-off between
growth and propagule production was studied in two undefended congeners with
different growth forms. Callyspongia armigera is a branching sponge, whereas
Callyspongia vaginalis occurs as a collection of tubes. C. armigera had higher growth
rates (0.36 ± 0.31 vs. 0.08 ± 0.11 % initial mass per day), higher number of attachment
points (2.31 ± 1.47 vs. 1.03 ± 0.18), and lower propagule production (0.04 ± 0.22 vs. 0.53
± 1.08 % area of reproductive propagules) compared to C. vaginalis. Branching sponges
can disperse by fragmentation, and therefore would allocate fewer resources to propagule
production. Results demonstrated a resource allocation trade-off between growth and
v
reproduction that is linked with morphology. To reduce complications from morphology-
linked resource allocation trade-offs, the remaining resource allocation trade-offs were
examined using only branching sponges.
Growth rates of undefended sponge species (Callyspongia armigera, Iotrochota
birotulata, and Niphates erecta) and defended species (Amphimedon compressa, Aplysina
fulva, Aplysina cauliformis and Ptilocaulis walpersi) were measured using predation
exclusion experiments. Growth was greater for undefended than defended sponges (0.89
± 0.01 vs. 0.77 ± 0.01 % g final g-1
initial day-1
). Winter growth was diminished in both
undefended and defended sponges compared to summer growth, but less so in
undefended sponges than defended sponges (significant season x defense interaction:
ANOVA, F = 10.01, df = 1, 1150, p = 0.002). Both comparative growth rates and
seasonal patterns of growth support a resource allocation trade-off between growth and
chemical defense among Caribbean coral reef sponges.
Reproductive output was quantified for six of the above branching species
(excluding Ptilocaulis walpersi), and in the tube sponge Callyspongia vaginalis.
Monthly samples were collected for a year, from which histological sections were made
for the quantification of reproductive propagules. Propagule production was highly
variable among the sponge species. On average, undefended and defended species had
the same reproductive output, and a resource allocation trade-off was not found between
reproduction and chemical defense. A simultaneous trade-off between growth and
propagule formation, such as the one between Callyspongia armigera and C. vaginalis,
could be confounding the pattern between propagule formation and chemical defense.
Finally, resource allocations to defense, growth and reproduction were consolidated to
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form a conceptual model of how resource allocation has influenced the evolution of
sponge communities on Caribbean coral reefs.
vii
ACKNOWLEDGEMENTS
None of this work would have been possible without the guidance, feedback and
support from my advisor Dr. Joseph Pawlik. My graduate committee, Dr. Richard
Dillaman and Dr. Stuart Borrett, provided many useful insights and suggestions. Mark
Gay taught me everything I know about histological procedures, and Dr. James Blum was
there to untangle my bouts of statistical confusion.
I am indebted to my lab mates for being my family away from home, and for all
their support and advice. Thanks especially to the new Dr. Timothy Henkel for always
being there to answer my questions about life, courses, procedures and statistics. Steven
McMurray, Tse-Lynn Loh, David Hines and Michael Echevarria were always happy to
bounce ideas off with me. I spent many long hours with the denizens of the Dillaman lab
- Carolina Priester, Anne Leaser, Ana Jimenez and Kristen Hardy, who were always a
source of encouragement and support. The office ladies, Tracie, Debby, Eleanor, Carol
and Lori saved me from being hopelessly entangled in bureaucracy.
There are many other folks I’ve met along the way, too many to list by name, who
have tirelessly answered my questions and provided technical support. These would
include the crew and participants on the R/V Seward Johnson, the folks at NURC, other
faculty and students in the department, and other people who work on sponges that I’ve
written to for advice and suggestions. Dr. Henry Feddern did an excellent job of sample
collection on my behalf.
My family has always been a silent but ceaseless fount of love and support for me,
and has given me the strength to be who I am.
viii
This research was funded by grants to Joseph R Pawlik from the National
Undersea Research Program at UNCW (NOAA NA96RU-0260) and from the National
Science Foundation, Biological Oceanography Program (OCE-0095724, 055468).
ix
LIST OF TABLES
Table Page
1. Details of growth experiments run at North Dry Rocks in Key Largo, Florida.
Species: ACO=Amphimedon compressa, ACA=Aplysina cauliformis,
AF=Aplysina fulva, CA=Callyspongia armigera, IB=Iotrochota birotulata,
NE=Niphates erecta, PW=Ptilocaulis walpersi; bold species are defended ...............34
2. Table 2: ANOVA results for factors affecting growth of sponges in Key Largo,
Florida. Significant factors are marked by an asterisk................................................34
x
LIST OF FIGURES
Figure Page
1. Abundance of Callyspongia armigera (CA) and Callyspongia vaginalis (CV)
from band quadrats along a line transect at North Dry Rocks reef, Key
Largo, Florida. Mean + SD. n = 10..............................................................................14
2. Average number of attachments per sponge for Callyspongia armigera (CA)
and Callyspongia vaginalis (CV) from band quadrats along a line transect at
North Dry Rocks reef, Key Largo, Florida. Mean + SD. n=101 and 119
respectively ..................................................................................................................14
3. Relative growth of Callyspongia armigera (CA) and Callyspongia vaginalis
(CV) in percent wet mass increase day-1
from eight caging experiments
conducted on reefs off Key Largo, Florida from 1996 to 2007. Mean + SD.
n=91 and 53 respectively. ............................................................................................15
4. Average reproductive output of Callyspongia armigera (CA) and
Callyspongia vaginalis (CV) from Conch Wall, Key Largo, Florida for
November 2007 to October 2008. Mean + SD. n=60.. ................................................15
5. Mean monthly reproductive output index (percent area reproductive
propagules) of Callyspongia armigera (CA) and Callyspongia vaginalis (CV)
from Conch Wall, Key Largo, Florida for November 2007 to October 2008.
Mean + SD. n=5...........................................................................................................16
6. Yearly increases in growth of Caribbean coral reef sponges in both caged and
uncaged treatments on reefs off Key Largo, Florida. Mean + SD; n in brackets.
Species with bold n are defended.................................................................................35
7. Daily growth in uncaged and caged treatments for undefended and defended
sponge species on reefs off Key Largo, Florida. Mean ± SE. N = 1158. ..................36
8. Daily growth for undefended and defended sponge species in different seasons
(summer/winter) on reefs off Key Largo, Florida. Mean ± SE. N = 1158 ..................36
9. Correlation between growth and defense.....................................................................37
10. Mean monthly ROI for seven coral reef sponge species in Key Largo, Florida.
n=5 ...............................................................................................................................55
11. Total yearly ROI for seven coral reef sponge species in Key Largo, Florida.
n=60, means + sd. Post-hoc comparisons were carried out using Wilcoxon’s
test with a Bon-ferroni correction. Different letter groups indicate a statistical
difference was found. Bold letters indicate defended species ....................................55
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12. Plot of growth, reproduction and defense. For each axis, the maximum value
occurring was set at 90 and all other values were scaled between 0-100.
Defended species (dots): Aplysina cauliformis, Aplysina fulva (solid dots),
Amphimedon compressa (hollow dot). Undefended sponges: Callyspongia
armigera (solid triangle), Callyspongia vaginalis (hollow triangle), Iotrochota
birotulata (solid square), Niphates erecta (hollow square). Theoretical
surface plot is overlaid, where the three axes sum to 100 (black mesh)......................56
13. Conceptual model of trade-offs between defense, growth and reproduction in
Caribbean coral reef sponges. ......................................................................................56
CHAPTER 1
FRAGMENTS VERSUS PROPAGULES: REPRODUCTIVE TRADE-OFFS FOR
TWO CALLYSPONGIA SPP. FROM FLORIDA CORAL REEFS
2
ABSTRACT
Fragmentation and propagule formation are reproductive strategies found in both
plants and animals, with the latter generally providing greater dispersal capability. When
both strategies occur, resource allocation theory predicts that growth and reproductive
resources should be divided between the two. On coral reefs, fragmentation of branching
corals and sponges allows for rapid habitat recolonization following disturbance by storm
events. In this study, we compared two congeneric sponges, Callyspongia armigera,
which grows in a branched form, and C. vaginalis, which does not, to test whether there
is a trade-off in growth or propagule formation for the two species. Both species were
common (10.1±3.7 vs. 11.9±3.0 per 100m2) and there was no significant difference in
their abundance on coral reefs off Key Largo, Florida. Growth rates (0.36±0.31 vs.
0.08±0.11 % initial mass day-1
) and the number of substratum attachment points
(2.31±1.47 vs. 1.03±0.18) were significantly higher for C. armigera compared to C.
vaginalis, but C. armigera produced less propagules than C. vaginalis (0.04±0.22 vs.
0.53±1.08 % area of reproductive propagules). Our results support a trade-off in growth
and reproductive strategies, suggesting that these closely related sponge species took
different evolutionary trajectories in reconciling their resource constraints.
3
INTRODUCTION
Trade-offs are implicated when two contrasting life history strategies co-exist
(Stearns 1992). For example, plants can either reproduce through propagule formation
(e.g. seeds) or by asexual fragmentation (e.g., rhizomes, runners, plantlets). Plants
allocate resources to physiological functions such as growth and reproduction from a
finite pool (Coley et al. 1985, Bazzaz et al. 1987). A trade-off arises when plants allocate
resources to vegetative growth for fragmentation instead of propagule formation.
Propagule formation provides several advantages. Propagules are smaller and
lighter than fragments and are able to disperse further (Gaylord et al. 2002). Also, most
propagules are sexual, which confers the advantages that sexual recombination provides –
it enables selection to break down negative gene combinations at different genetic loci,
and increases genetic diversity (Hoekstra 2005, Charlesworth 2007). Yet, fragmentation
provides benefits too. By investing in vegetative growth, organisms can rapidly increase
in biomass to colonize new areas (Abrahamson 1975). By breaking up plants into
independent units, fragmentation also reduces the spread of infection between clones
(Hay and Kelly 2008).
For plants located in or near water bodies, fragments can be carried by water to a
new location and regenerate to form new ramets (physiologically independent clones). In
British riverine plants, there was a clear trade-off in allocation to dispersal mode between
Sparganium emersum and Ranunculus trichophyllus where fragments survive and take
root successfully, and Luronium natans, Hippuris vulgaris and Elodea canadensis that
produce propagules for dispersal (Barrat-Segretain 1996, Barrat-Segretain et al. 1998).
Fragments of the seagrasses Halodule wrightii and Halophila johnsonii are able to take
4
root but the former is viable for longer periods of times and can disperse over longer
distances (Hall et al. 2006).
Vegetative fragmentation is responsible for the success of many invasive aquatic
plants. Invasive riverine vegetation such as Mimulus guttatus (Truscott et al. 2006)
employ fragmentation and recolonisation to rapidly spread downstream after
unpredictable flood pulses. Rapid colonisation by fragments also explains the success of
the invasive seagrass Posidonia oceanica (Di Carlo et al. 2005) and the seaweed
Caulerpa taxifolia compared to the local species C. prolifera and C. verticillata (Smith
and Walters 1999) in the Mediterranean.
Among animals, simple clonal organisms (cnidarians) or those with indeterminate
integration of body plan (sponges) also adopt fragmentation as a strategy for dispersal
(Tunnicliffe 1981, Lasker 1984, Wulff 1991). Fragmentation provides benefits to corals
and sponges that grow on coral reefs. Coral fragments exhibit higher survivorship due to
their larger size compared to recruits and juveniles (Highsmith 1982). Corals and
sponges also are able to recover more quickly after disturbances such as storm damage as
a result of fragmentation and subsequent reattachment (Highsmith 1982, Wulff 1995).
Since most coral diseases are spread by contact with infected tissue, Highsmith (1982)
also proposed that fragmentation may be a way for colonies to limit the spread of disease.
Clonal growth is very successful as a strategy and the use of small coral fragments has
been proposed as a method for repopulating coral reefs (Shafir et al. 2001).
Sponges are dominant members of the benthic sessile community on Caribbean
coral reefs (Targett and Schmahl 1984, Aronson et al. 2002, Maliao et al. 2008). Yet,
there have been a limited number of studies addressing dispersal trade-offs in sponges,
5
particularly the idea that the “branching” morphology maximizes fitness by enhancing
the exploitation of fragmentation. Erect branching sponges have been shown to
recolonize an area of cleared substrate quickly (Wulff 1991, Wulff 1995). Tsurumi and
Reiswig (1997) noted that the production of sexual propagules was very infrequent in
Aplysina cauliformis, a thin branching sponge. They suggested that the thin branching
morphology may be an adaptation for fragmentation, but they did not compare
reproduction in A. cauliformis with reproduction in non-branching congeners.
Callyspongia armigera and C. vaginalis are two sponges that are very common
on Caribbean reefs. The former consistently grows as a thin or branching rope, which is
more suited for fragmentation. The latter, also known as the common vase sponge,
grows in clusters of tubes. Both species produce propagules that are assumed to be
sexual products, but sperm have never been observed. The lack of sperm is most likely
due to sampling bias. To determine whether there is a resource trade-off for these two
species with contrasting dispersal strategies, we examined rates of growth and propagule
formation for each. We also examined the number of points of attachments for each
species to compare how easily the sponge species can reattach to the substratum. A
species that fragments would be expected to have more points of attachment, higher rates
of vegetative growth, and lower production of propagules.
METHODS AND MATERIALS
Collections and experiments were conducted at North Dry Rocks (N25o07.850’
W80o17.521’), Conch Wall (N24
o56.440’ W80
o27.230’), and Carysfort Reef
(N25o12.860’ W80
o12.810’) off Key Largo in Florida. North Dry Rocks and Carysfort
6
Reef are shallow patch reefs at 8m depth. The Conch Wall site was located on the
shallow reef flat, approximately 12m in depth, and contained similar assemblages of
sponge species. For all sites, the predominant substratum was limestone coral pavement,
which was interspersed with small patches of overlying sand. These sites were chosen
because both study species occurred there.
To document relative abundances of the two sponge species, a survey was
conducted at North Dry Rocks to determine relative abundances of both species on the
reef. Ten 20m x 5m band quadrats were surveyed along a continuous transect line and
the number of sponge individuals of each species that lay within the band was recorded.
Sponges that grew as a solitary mass were counted as individuals.
From the same quadrats, the number of points in which each sponge was attached
to the substratum were counted. Examples of substratum included surfaces that provided
a firm anchor, such as the surrounding limestone, and on other organisms, such as
gorgonians, that were firmly anchored to the limestone.
Growth data were obtained from eight predation exclusion experiments conducted
between 1996 and 2007. The experimental start dates were 6 May 1996, 19 May 1997,
12 May 1999, 6 May 2000, 7 May 2002, 5 June 2003, 25 May 2006, 4 June 2007. Each
experiment lasted 124 to 176 days. Cages were constructed with 1-inch vexar and cable
ties and secured on the reef with nails. Sponges were carefully collected from the
surrounding reef, weighed on an electronic scale, tagged, returned to the same reef and
secured to the surfaces of bricks inside cages. At the end of the experiment, sponges
were retrieved and weighed in the same way. Growth rates were measured as a change in
7
mass presented as a percentage of initial mass, and corrected for duration by dividing
over the number of days the sponges were left on the reef.
To determine reproductive output, five sponges of each species were collected
monthly from Conch Wall in Key Largo from November 2007 to October 2008, and
processed for histology. Three 1cm cubes were punched either out of the wall of the tube,
or collected along the length of the sponge for each specimen. The sponge cubes were
immediately fixed in 10% formalin buffered with sea water. Specimens were then rinsed
with buffer and deionised water, dehydrated in a series of ethanol, and embedded in
paraffin using toluene as a clearing agent. Using a rotary microtome, 10µm sections were
made and stained with haematoxylin and eosin. Specimens were then viewed with an
Olympus BX60 microscope with a SPOT camera attached. A total of twenty views of
each specimen were haphazardly photographed at 4x magnification to give a total
scanned area of 128.92mm2
for each sponge. The surface area of any propagules present
was quantified using the image analysis software ImageJ (Rasband 1997). Surface area
measurements were then converted to a percentage of the total surface area, which has
been termed the “Reproductive Output Index” (ROI), enabling reproduction to be
compared between species (Whalan et al. 2007).
One- and two-tailed Student’s t-tests were used to compare survey and growth
experiment findings, and the non-parametric alternative, Wilcoxon’s rank sum, was used
for the reproduction data due to the lack of normality in the data.
8
RESULTS
In surveys on shallow reefs off Key Largo, Florida, equal abundances were found
for both Callyspongia armigera and C. vaginalis, at 11.9 and 10.1 per quadrat
respectively (Two-tailed t-test, t=2.101, df=18, p=0.2471; Fig. 1). Callyspongia
armigera had an average of 2.22 attachments per individual, which was significantly
more than 1.03 attachments per individual for C. vaginalis (One-tailed t-test, t=-8.417,
df=18, p<0.0001; Fig. 2).
Sponge growth in cages was over four times higher for C. armigera (n=93,
mean=0.358 % initial mass day-1
) compared to C. vaginalis (n=53, mean=0.079 % intial
mass day-1
; one-tailed t-test, t=6.395, p<0.0001; Fig. 3).
Both C. armigera and C. vaginalis brood their propagules (larvae or oocytes, see
discussion) in distinct chambers. Propagules appear identical for both species, are 0.5mm
in length and can be easily seen without magnification. When reproductive propagules
were present, the ROI of each individual was comparable between species (1.22 for C.
armigera and 1.78 for C. vaginalis). Propagules are of similar size and appearance in
both species.
After monthly samples for one year (N=60), only two reproductive individuals of
C. armigera were found, one in March and another in October 2008 (Fig. 5). On the
other hand, 18 of 60 samples (30%) of C. vaginalis exhibited propagules, and there was
seasonality in their production, with propagules found in December 2007, and from May
to September 2008 (Fig. 5). No sperm were observed in the samples. Overall, C.
armigera had a much lower total annual ROI than C. vaginalis (Wilcoxon’s rank sum test,
χ2=15.317, p<0.0001; Fig. 4).
9
DISCUSSION
As for some aquatic plants (Barrat-Segretain 1996, Barrat-Segretain et al. 1998,
Hall et al. 2006), there seems to be a clear trade-off between investment in fragmentation
versus propagule formation for Callyspongia armigera and C. vaginalis, both of which
are equally abundant on reefs off Key Largo, Florida. More points of attachment, higher
growth rates and lower reproduction are consistent with the hypothesis that the branching
growth form of C. armigera disperses mainly by fragmentation. Via fragmentation, C.
armigera can quickly colonize free substratum after disturbances such as hurricanes,
which occur frequently on the Florida reef tract.
Branching sponge tissue does not have lower tensile strength than the tissue of
non-branching species when comparing tissue strips with similar cross-sectional surface
areas (Chanas and Pawlik 1995). However, whether branching species may be more
susceptible to fragmentation on the whole as a colony remains to be tested. Branching
sponge species often extend off the substratum, presenting a greater surface area to
current and wave action. Fish predation would also be more likely to generate fragments
off a branching sponge since any bites would separate or weaken sections of the sponge.
Sponges are not the only benthic sessile invertebrates where branching growth
forms are thought to be advantageous in fragmentation. Investment in propagules is also
reduced in arborescent and vine-like bryozoans which disperse via fragmentation
(Thomsen and Hakansson 1995). The gorgonian Plexaura sp. (Lasker 1984) and
staghorn coral Acropora cervicornis (Tunnicliffe 1981) also disperse mainly by clonal
fragmentation, with little evidence of sexual propagule formation. High levels of asexual
10
reproduction have also been reported in the branching coral Pocillopora damicornis
(Sherman et al. 2006) and other branching reef-building corals on the Great Barrier Reef
(Ayre and Hughes 2000), which is consistent with fragmentation being the dominant
mode of reproduction in such branching forms.
Clonal (asexual) fragmentation is often cited as an evolutionary alternative to
sexual reproduction (Tunnicliffe 1981, Lasker 1984, Wulff 1991, Thomsen and
Hakansson 1995, Ayre and Hughes 2000, Sherman et al. 2006). Vegetative growth
would be favored for fragmentation whereas brooding propagules would be favored for
sexual recombination. However, asexual reproduction is usually favored in stable,
unchanging environments (Abrahamson 1975, Silvertown 2008), contrary to what would
be expected if fragmentation is more successful for recovery after disturbance. Hence,
fragmentation would be expected to produce the lowest genotypic diversity at
intermediate levels of disturbance (Coffroth and Lasker 1998). At low disturbance,
fragmentation would not occur. At high disturbance, organisms reproduce sexually,
creating genotypic diversity to cope with environmental fluctuations.
While Callyspongia vaginalis is ubiquitous, C. armigera does not occur on all
reefs in the Florida Keys, but can only be found in sites like North Dry Rocks and Pickles
Reef. It occurs in comparable abundance where it is found. The patchy distribution of C.
armigera may be due to differences in disturbance, flow regimes, or habitat complexity
between reefs. Range and mode of dispersal is a key difference between fragments and
propagules. A combination of flow rate and habitat complexity affects the dispersal of
fragments and therefore may explain the distribution of C. armigera. Low flow would
limit the dispersal of fragments which are heavier and less buoyant than propagules, and
11
high flow may flush out fragments before they can attach. Habitat complexity provides
microhabitats where fragments may settle for long enough to attach.
One interesting finding was that no sperm were observed in histological samples
of either sponge species, making it impossible to determine if propagules are sexual or
asexual. Sperm have been absent in other studies of sponge reproduction (Fell 1989,
Corriero et al. 1996), while in others, heavily skewed sex ratios for both females
(Tsurumi and Reiswig 1997, Mercurio et al. 2007) and males (Whalan et al. 2007) have
been reported. The most parsimonious explanation for the absence of sperm despite the
presence of propagules within the brood chambers is that the incubation time for sperm is
so short that a monthly sampling scheme would overlook them (Mercurio et al 2007).
While the high variation in sponge sex ratios is fascinating, it does not detract from the
fact that significantly fewer Callyspongia armigera contained propagules compared to C.
vaginalis using the same sampling methodology.
Callyspongia armigera and C. vaginalis are not the only Callyspongia species
found on reefs in the Florida Keys. Callyspongia fallax and C. plicifera also occur, but in
much lower abundances. Although reproductive data is lacking for C. fallax and C.
plicifera, they do not appear to be as fecund as C. vaginalis (Pawlik, pers. obs.), and like
C. vaginalis, they are both vase sponges that do not fragment like C. armigera. The low
fecundity of C. fallax and C. plicifera may explain their low abundance on the reef.
In summary, this study has provided the first comparative evidence that there is a
trade-off between reproductive modes and dispersal strategies in two species of closely
related coral reef sponges. The trade-off is related to morphology, with the branching
12
species exhibiting traits associated with higher fragmentation compared to the tube
species.
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207-226.
Hoekstra, R. F. 2005. Evolutionary biology: why sex is good. - Nature 434: 571-573.
Lasker, H. R. 1984. Asexual reproduction, fragmentation, and skeletal morphology of a
plexaurid gorgonian. - Mar Ecol Prog Ser 19: 261-268.
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cydonium (Jameson 1811) (Porifera, Demospongiae) from a semi-enclosed
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14
Targett, N. M. and Schmahl, G. 1984. Chemical ecology and distribution of sponges in
the Salt River Canyon, St. - Croix, USVI USA: NOAA Technical memorandum OAR
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15
0
2
4
6
8
10
12
14
16
CA CV
Ab
un
dan
ce (
per
100m
2)
n=10
Fig. 1: Abundance of Callyspongia armigera (CA) and Callyspongia vaginalis (CV)
from band quadrats along a line transect at North Dry Rocks reef, Key Largo, Florida.
Mean + SD. n=10.
0
0.5
1
1.5
2
2.5
3
3.5
4
CA CV
Avera
ge a
ttach
men
ts p
er
ind
ivid
ual
n=119n=101
*
Fig. 2: Average number of attachments per sponge for Callyspongia armigera (CA) and
Callyspongia vaginalis (CV) from band quadrats along a line transect at North Dry Rocks
reef, Key Largo, Florida. Mean + SD. n=101 and 119 respectively.
16
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
CA CV
Avera
ge g
row
th r
ate
(%
in
itia
l m
ass p
er
day)
n=91 n=53
*
Fig. 3: Relative growth of Callyspongia armigera (CA) and Callyspongia vaginalis (CV)
in percent wet mass increase day-1
from eight caging experiments conducted on reefs off
Key Largo, Florida from 1996 to 2007. Mean + SD. n=91 and 53 respectively.
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
1.8
CA CV
Avera
ge R
OI
n=60 n=60
*
Fig. 4: Average reproductive output of Callyspongia armigera and Callyspongia
vaginalis from Conch Wall, Key Largo, Florida for November 2007 to October 2008.
Mean + SD. n=60.
17
0
0.5
1
1.5
2
2.5
3
3.5
Nov-
07
Dec-
07
Jan-
08
Feb-
08
Mar-
08
Apr-
08
May-
08
Jun-
08
Jul-
08
Aug-
08
Sep-
08
Oct-
08
RO
I
CA CV
n=5
Fig. 5: Mean monthly reproductive output index (percent area reproductive propagules)
of Callyspongia armigera (CA) and Callyspongia vaginalis (CV) from Conch Wall, Key
Largo, Florida for November 2007 to October 2008. Mean + SD. n=5.
CHAPTER 2:
IS THERE A TRADE-OFF BETWEEN GROWTH AND DEFENSE AMONG
CARIBBEAN CORAL REEF SPONGES?
19
ABSTRACT
Like all organisms, sponges allocate resources to life functions such as growth
and reproduction. Additionally, some sponges may produce defensive compounds in
order to deter predation. Assuming resources are limiting, species that produce defensive
metabolites would be expected to allocate fewer resources to growth and reproduction.
To examine trade-offs between chemical defense and growth, predator exclusion
experiments were conducted to compare the growth rates of seven common Caribbean
sponge species with branching morphology: the undefended species Callyspongia
armigera, Iotrochota birotulata, and Niphates erecta, and defended species Amphimedon
compressa, Aplysina cauliformis, Aplysina fulva, and Ptilocaulis walpersi. A three-factor
ANOVA was used to compare the effects of chemical defense (undefended/defended),
treatment (uncaged/caged) and season (summer/winter) on growth. Overall, growth was
greater for undefended than defended sponges (0.89 ± 0.01 vs. 0.77 ± 0.01 % g final g-1
initial day-1
). Winter growth was diminished in both undefended and defended sponges
compared to summer growth, but less so in undefended sponges than defended sponges
(significant season x defense interaction: ANOVA, F = 10.01, df = 1, 1150, p = 0.002).
Growth rates and seasonal growth patterns show sponges use different allocation patterns
to cope with resource constraints.
20
INTRODUCTION
Terrestrial plants produce many chemical and physical defenses against herbivory
(Berenbaum and Zangerl 2008). Yet, under similar levels of herbivory, defended plants
commonly co-occur with undefended plants. Several hypotheses have been put forth to
explain phenotypic, genetic, and geographical variation in plant defenses (Stamp 2003,
Agrawal 2007). According to the resource availability hypothesis, plants allocate
resources from a finite pool, resulting in trade-offs between defense, growth and
reproduction (Coley et al. 1985, Bazzaz et al. 1987, Bazzaz and Grace 1997). A plant
can either resist predation by producing deterrent compounds or tolerate predation by
allocating resources to growth and reproduction, and many examples of growth and
defense trade-offs have been described (Herms and Mattson 1992, Fine et al. 2006).
Like terrestrial plants, Caribbean coral reef sponges are also subject to grazing
(Randall and Hartman 1968, Dunlap and Pawlik 1996, Pawlik 1998, León and Bjorndal
2002). Organic extracts of the tissues of 73 Caribbean sponge species exhibited a wide
range of feeding deterrent activities in experiments with the blue-head wrasse
Thalassoma bifasciatum (Pawlik et al. 1995). Based on laboratory and field feeding
experiments, as well as predation exclusion experiments, (Pawlik 1998) grouped sponges
into three categories –“preferred” sponge species are rapidly grazed down and only
survive in cryptic refugia, while “undefended” and “defended” species both co-exist on
the reef. The defended species are avoided by fish predators while undefended species
are consumed by sponge eating fishes. Some of the secondary metabolites responsible
for deterrent activity in defended sponges have been isolated (Albrizio et al. 1995,
Puyana et al. 2003, Nuñez et al. 2008). Assuming these complex compounds require
21
metabolic energy to synthesize, store and elaborate, it should be expected that there is a
trade-off between growth and chemical defenses in sponges.
There is some evidence for a growth-defense trade-off in sponges. Hoppe (1988)
found variability between predation deterrence, growth and healing in three defended
sponges Neofibularia nolitangere, Ircinia strobilina and Agelas clathrodes, and
concluded that the variability may be due to differences in resource allocation strategies.
Walters and Pawlik (2005) investigated wound healing in ten species of Caribbean coral
reef sponges, and found that undefended species had faster rates of wound healing than
defended species. Wound healing occurs after sponge tissues are damaged, and it
proceeds at a much faster rate than regular somatic growth (Ayling 1983). In sponge
species that are grazed often by reef organisms, rapid wound healing should occur to
repair damaged tissue and prevent microbial colonization and necrosis (Ayling 1983,
Walters and Pawlik 2005).
More recently, Pawlik et al. (2008) examined patterns of colonization on a large
shipwreck off Key Largo, Florida, 4.5 years after it was sunk to form an artificial reef.
Undefended sponge species predominated on the surface of the wreck in terms of size
and abundance compared to the sponge community of surrounding reefs. Small
individuals of the most common defended sponge species were only found in a
subsequent survey 1.5 years later. It was concluded that more rapid growth or faster
recruitment of the undefended sponge species was consistent with a trade-off between
chemical defense and growth or reproduction.
To more directly investigate trade-offs between growth and chemical defense in
undefended and defended sponge species, predator exclusion experiments were
22
performed using seven species of branching sponges that commonly occur on Caribbean
coral reefs. Branching species were easier to transplant and secure than species with
other growth morphologies, and had higher survival rates after transplantation. The
undefended species Callyspongia armigera, Iotrochota birotulata, and Niphates erecta;
and defended species Amphimedon compressa, Aplysina cauliformis, Aplysina fulva, and
Ptilocaulis walpersi were chosen for this experiment, because these are the most
abundant branching sponges on Caribbean reefs (Engel and Pawlik 2005).
METHODS AND MATERIALS
The growth study was conducted on North Dry Rocks reef (N25o07.850’
W80o17.521’) in Key Largo, Florida. Cages measuring 30cm by 30cm by 30cm were
constructed with 1-inch vexar and cable ties and secured on available patches of the
limestone substratum with nails and fasteners. Each cage shared a base with an uncaged
sponge which served as the uncaged controls.
Sponges were collected from the surrounding reef at North Dry Rocks. Pieces of
sponge ~10cm in length were carefully cut with a sharp razor and kept immersed in fresh
seawater while transported to the laboratory where they were quickly tagged and weighed
using an electronic scale. Sponges were kept submerged to avoid air bubble formation in
their tissues. Within a few hours, sponges were transported back to the same reef where
they were collected and secured on bricks inside and outside of cages. At the end of 5-6
months, sponges were retrieved and transported back to the laboratory, where they were
kept submerged while cleaned of any fouling organisms before being weighed in the
same way as before.
23
Nine successful iterations of the experiment were conducted between the years
1999 and 2008, with 1-4 sponge species (20 cages and 20 uncaged controls per species)
used in each iteration. Of the nine iterations, six were conducted in the summer, and
three in the winter. Winter runs were added later, starting in 2005. Details are
summarized in Table 1.
For each sponge, a growth index (% gfinal g-1
initial day-1
) was calculated as follows:
growth = final mass / initial mass / number of days in the iteration x 100%. This index
was comparable across all iterations and species and is the unit of measure used in the
statistical analyses. For easier comparisons with existing literature, change in mass was
also calculated as: (final mass – initial mass) / initial mass / number of days in the
iteration x 100%. Percentage mass increase was then multiplied by 365 to give yearly
growth rates for each species (% growth year-1
, Fig. 1).
Differences in growth rates were analyzed using an ANOVA (Mixed procedure in
SAS 9.1.3 (S.A.S. 2005)) with defense (undefended/defended), treatment (uncaged/caged)
and season (summer/winter) as the factors. Year, pair number and individual sponge
species were also initially included as random factors, but did not have significant effects
on the model, and so were excluded. Sponges that were missing at the final collection
were excluded from the analysis because it was impossible to tell if the sponges had died
or were improperly secured and swept away.
RESULTS
Growth was highly variable in all sponge species (Fig. 6). All species exhibited
overall positive growth except for uncaged treatments of the defended sponge Ptilocaulis
24
walpersi. Growth rates of caged sponges ranged from 0 to 133 % growth year-1
.
Uncaged species had slightly lower growth rates, at -1 to 105 % growth year-1
.
There was a significant interaction between treatment and defense (ANOVA, F =
5.53, df = 1, 1150, p = 0.0189; Table 2; Fig. 7). In the undefended sponge species, the
caged treatment exhibited significantly higher growth than the uncaged treatment (0.93 ±
0.01 [mean ± standard error] vs. 0.86 ± 0.01 % gfinal g-1
initial day-1
; t = 3.94, df = 1150, p <
0.0001). There was no difference in growth between caged and uncaged treatments in the
defended sponge species (0.78 ± 0.01 vs. 0.77 ± 0.01 % gfinal g-1
initial day-1
; t = 0.42, df =
1150, p = 0.672). In both caged and uncaged treatments, undefended sponge species
exhibited higher growth than defended species. Comparing growth overall, undefended
species had higher growth than defended species (0.89 ± 0.01 vs. 0.77 ± 0.01 % gfinal
g-1
initial day-1
).
There was also a significant interaction between season and defense (ANOVA, F
= 10.01, df = 1, 1150, p = 0.002; Table 2; Fig. 8). Growth occurred in both summer and
winter, but winter growth rates were lower than summer growth rates for both
undefended and defended sponge species. The defended and undefended species differ in
the extent to which growth is reduced in winter. In the undefended species, winter
growth was marginally lower than summer growth (0.87 ± 0.02 vs. 0.92 ± 0.01 % gfinal
g-1
initial day-1
; t = 2.36, df = 1150, p = 0.018). In the defended species, winter growth was
much lower than summer growth (0.71 ± 0.02 vs. 0.84 ± 0.01 % gfinal g-1
initial day-1
; t =
6.46, df = 1150, p < 0.0001).
25
DISCUSSION
Growth and chemical defense
From the comparison of seven sponge species, results demonstrate a trade-off
between growth and defense in Caribbean coral reef sponges. The growth and defense
trade-off, combined with other evidence (Walters and Pawlik 2005, Pawlik et al. 2008),
provides support for the resource availability hypothesis in a completely different group
of organisms from the terrestrial plants (Coley et al. 1985, Bazzaz et al. 1987, Bazzaz and
Grace 1997). Growth was greater in undefended sponge species that tolerate rather than
resist predation. Growth, like wound healing, should be negatively correlated with
defense, but continuous investment of resources in growth is different from faster wound
healing in response to predation by fishes, which is a response triggered by tissue damage
(Ayling 1983). Undefended sponge species are not only able to respond more quickly
after predation to regenerate tissue lost (Walters and Pawlik 2005), but they also invest
more in growth that occurs independent of tissue damage.
To evaluate the relationship between growth rate and chemical defense, growth
rates were plotted against palatability (values obtained from Pawlik et al. 1995) to
generate a figure similar to that reported in Walters and Pawlik (2005) (Fig. 9). Pawlik et
al. (1995) tested palatability of crude organic extracts of sponge tissue using as a scale the
number of extract-treated pellets eaten out of ten, with ten being completely palatable,
and zero being completely deterrent. Palatability was correlated with growth rates for the
sponge species used in this study. The overall trend of positive correlation between
growth rates and palatability was the same as that for wound healing, but growth (r2 =
0.20) was more poorly correlated with palatability than wound healing (r2 = 0.64).
26
Regular growth is not a response to tissue damage, unlike wound healing. Furthermore,
during normal growth, resources can be allocated to remodeling the tissue matrix in ways
that do not result in an overall increase in mass. The low r2 value suggests that defense
allocation does not account for much of the variability in growth between the different
species. This is not surprising because other resource allocation trade-offs are occurring
(e.g. a trade-off between defense and reproduction), and so the pattern between any single
trade-off becomes less clear (Mole 1994). This growth study was limited to branching
sponges because they are less likely to divert resources to propagule formation (Leong
and Pawlik, in prep). Hence, an examination of the trade-off between chemical defense
and reproduction would be needed to obtain a more complete picture.
In the Caribbean, coral reef sponges are eaten primarily by fishes and turtles
(Randall and Hartman 1968, Dunlap and Pawlik 1996, Leon and Bjorndal 2002).
Observations of fish feeding on reef sponges show spongivorous fishes selectively
feeding on undefended sponges (Dunlap and Pawlik 1996). In this experiment,
chemically undefended sponge species actually grew at the same rate in both the caged
and uncaged treatments, but the uncaged sponges appeared to grow slower because
selective grazing concurrently decreased the mass of the uncaged undefended sponges.
Defended sponge species had the same growth rates in both the caged and uncaged
treatments. Selective predation on undefended species that co-exist with defended
species has been recorded in-situ for plants and lichens (Coley 1983, Westerbergh and
Nyberg 1995, Nimis and Skert 2006), but not for any of the other sponge communities
where both defended and undefended species co-exist (Van de Vyver et al. 1990, Uriz et
al. 1991, Burns and Ilan 2003, McClintock et al. 2005). Selective predation on
27
undefended species, which allocate more resources to growth, explains how both
chemically defended and undefended sponges can occur on the same coral reef.
Of the sponge species tested for this study, Callyspongia armigera, Iotrochota
birotulata and Niphates erecta lack anti-predatory chemical extracts. Amphimedon
compressa produces a pyridinium alkaloid that is highly deterrent to predators (Albrizio
et al. 1995), Aplysina spp. brominated tyrosine derivatives similar to all members of the
Verongidae (Puyana et al. 2003, Nuñez et al. 2008). Like members of the potently
defended genus Agelas, Ptilocaulis walpersi contains bromopyrroles and oroidin-class
metabolites (Wright et al. 1991). The synthetic pathways and costs of sponge secondary
metabolites are not well understood, but secondary metabolites are expected to be costly
due to the requirement for raw materials, the production and storage of metabolites, and
prevention of autotoxicity (Van Alstyne et al. 2001). Nevertheless, some sponge species
have also been known to gain the advantages of chemical defenses produced by their
symbionts at little to no direct cost (Haygood et al. 1999). The existence of a trade-off in
growth rates between defended and undefended sponge species demonstrated herein
suggests that chemical defenses for some Caribbean coral reef sponges may incur a cost.
Unlike the situation with terrestrial plants, predation is generally unaffected by
physical defenses or nutritional quality in sponges (McClintock 1987, Chanas and Pawlik
1995, Chanas and Pawlik 1996). While sponge spicules, which are often sharp glass
shards, have been observed to have effects in feeding assays in some experiments (Burns
and Ilan 2003, Hill et al. 2005), more rigorous treatments of their interaction with
chemical defenses provide little evidence of a defensive role (Jones et al. 2005), perhaps
because most coral reef fishes have mouth parts that are designed for processing hard
28
parts. Additionally, the issue of whether defenses are optimized in Caribbean sponges
remains contested. Optimization could occur via activation or induction of defenses, a
situation that has been demonstrated in some terrestrial plants and algae in which
bioactive compounds are rapidly converted from inactive precursors, or produced only in
response to predation (Cronin and Hay 1996, Agrawal 1998, Harvell 1990). Thus far,
there is little evidence of optimization in Caribbean sponges (Chanas and Pawlik 1997,
Swearingen and Pawlik 1998, Puyana et al. 2003). Spatial and temporal fluctuations in
defense, such as those in the Mediterranean sponge Crambe crambe may also indicate
optimization of defense. Uriz et al. (1995) found greater allocation to mineral and
organic structures and lower allocation to reproduction in shaded compared to light
habitats. They attributed differences in allocation to defensive structures among Crambe
crambe in light and shaded habitats to variation in resource availability or competitive
pressure between the two habitats. Crambe crambe also regulates its production of
defensive chemicals according to size, season, and environmental factors, optimizing the
use of available resources (Becerro et al., 1997). However, no such temporal fluctuations
have been found in the Caribbean sponges.
Growth rates and seasonality
Growth occurred for all sponge species except for the uncaged treatment of
Ptilocaulis walpersi, a chemically defended species. Negative growth, or shrinkage, has
been described for other sponge species (Elvin 1976, Hoppe 1988, Garrabou and Zabala
2001, McMurray et al. 2008). Growth rates in the present study should be robust,
because they were averaged over a five month period, sufficient for recovery and growth
in branching sponge species.
29
Growth rates of uncaged sponge species, subject to tissue loss from predation, of -
1 to 105% are comparable to rates reported in the literature. In comparison, Xestospongia
muta, the giant barrel sponge, increased in volume at 52% per year (McMurray et al.
2008). Hoppe (1988) found growth rates of 7 to 19% per year for Neofibularia
nolitangere, Ircinia strobilina and Agelas clathrodes, and (Reiswig 1973) reported
growth rates of 5 to 60% per year for Mycale sp., Verongula gigantea and Tethya crypta
in Jamaica. Our results reveal that some rope sponges have very high growth rates, with
Aplysina fulva and Callyspongia armigera able to double their mass per year.
Sponge growth was higher in summer than in winter. Different rates of growth in
sponges have been attributed to both physical factors (temperature, environmental stress,
water flow and depth) and physiological factors related to resource allocation trade-offs
(e.g. seasonal reductions in growth due to investment in reproduction). For the most part,
physical factors that affect growth are linked to food availability and delivery, with
higher growth occurring in the warmer months when more food is available (Elvin 1976,
Duckworth et al. 2004, McMurray et al. 2008). Barthel (1986) did not find any
correlations between temperature and food availability in the Baltic Sea, and suggested
that temperature may be linked with respiration rates to explain growth patterns.
However, conditions in the Baltic Sea are very different from conditions in the Caribbean.
Depth associated picoplankton availability has been found to affect growth in some
species (Lesser 2006, Trussell et al. 2006) but not others (McMurray et al. 2008).
Duckworth et al. (2004) found that sponges grew fastest in areas of high flow and
postulated that this may be due to improved delivery of food, which enables sponges to
feed with minimal pumping. Verdenal and Vacelet (1990) found that increased turbidity
30
decreased growth in sponges, and suggested that this may be due to clogging of sponge
pores that obstruct feeding. Seasonal growth can also be decreased due to negative
correlation with reproductive input (Turon et al. 1998) or conversion of feeding
choanocytes to sperm cells (Duckworth et al. 2004).
In Caribbean coral reef sponges, higher growth in the warm summer months
(May-October) corresponded with higher rates of reproduction (Leong, unpublished data).
If resources are more limited in winter, then defended sponge species should grow less
during the winter compared to undefended sponge species, when allocation to defense
may consume a greater proportion of overall resources available assuming that metabolite
production remains the same in both seasons. Such a relationship was found in the data
in the form of a significant interaction between season and defense. Hence, seasonal
growth patterns were also consistent with the resource allocation hypothesis.
In summary, trade-offs between chemical defenses, growth rates and seasonal
growth patterns support the resource allocation hypothesis in some species of Caribbean
sponges. Although a trade-off was found, a low correlation between growth and defense
indicates other resource trade-offs may be obscuring the relationship. The trade-off
between reproductive input and defense will be examined in a paper to follow.
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Swearingen III DC, Pawlik JR (1998) Variability in the chemical defense of the sponge
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35
Table 1: Details of growth experiments run at North Dry Rocks in Key Largo, FL.
Species: ACO=Amphimedon compressa, ACA=Aplysina cauliformis, AF=Aplysina fulva,
CA=Callyspongia armigera, IB=Iotrochota birotulata, NE=Niphates erecta,
PW=Ptilocaulis walpersi; bold species are defended.
Year Season Start date End date Duration (days) Species
2007 Winter 29-Nov-07 07-May-08 160 CA, IB, ACO, ACA
2007 Summer 04-Jun-07 28-Nov-07 176 CA, IB, ACO, ACA
2006 Winter 15-Nov-06 29-May-06 195 CA, IB, ACO, ACA
2006 Summer 25-May-06 12-Nov-06 171 CA, IB, ACO, ACA
2005 Winter 12-Dec-05 23-May-05 162 CA, ACA
2003 Summer 05-Jun-03 06-Oct-03 124 CA, IB, NE, AF
2002 Summer 07-May-02 14-Oct-02 159 CA, NE, AF
2000 Summer 06-May-00 03-Oct-00 151 CA, IB, ACA, PW
1999 Summer 12-May-99 05-Oct-99 147 IB
Table 2: ANOVA results. Significant factors are marked by an asterisk.
Source df F p
Season (Winter/Summer) 1,1150 40.42 <0.0001* Defense (Defended/Undefended) 1,1150 76.33 <0.0001* Treatment (Caged/Uncaged) 1,1150 8.86 0.0030* Season x Defense 1,1150 10.01 0.0016* (Fig. 8) Season x Treatment 1,1150 1.05 0.3051 Defense x Treatment 1,1150 5.53 0.0189* (Fig. 7) Season x Defense x Treatment 1,1150 0.00 0.9997
Fig. 6: Yearly increases in growth of Caribbean coral reef sponges in both caged and uncaged treatments on reefs off Key Largo,
Florida. Mean + SD; n in brackets. Species with bold n are defended.
-50
0
50
100
150
200
250
Iotrochota
birotulata
Niphates erecta Callyspongia
armigera
Amphimedon
compressa
Aplysina
cauliformis
Aplysina fulva Ptilocaulis
walpersi
% g
row
th p
er
year
Uncaged Caged
(137, 137) (40, 40) (153, 155) (75, 72) (115, 115) (40, 40) (20, 19)
0.005
0.105
0.205
0.305
0.405
0.505
0.605
0.705
0.805
0.905
1.005
Uncaged Caged
Gro
wth
per
day
(% g
fin
al
g-1
in
itia
l d
ay
-1)
Defended Undefended N=1158
Fig. 7: Daily growth in uncaged and caged treatments for undefended and defended
sponge species on reefs off Key Largo, Florida. Mean ± SE. N = 1158.
0.005
0.105
0.205
0.305
0.405
0.505
0.605
0.705
0.805
0.905
1.005
Undefended Defended
Gro
wth
per
day
(% g
fin
al
g-1
in
itia
l d
ay
-1)
Summer Winter N=1158
Fig. 8: Daily growth for undefended and defended sponge species in different seasons
(summer/winter) on reefs off Key Largo, Florida. Mean ± SE. N = 1158.
38
y = 4.0122x + 56.515
r2 = 0.2001
0
20
40
60
80
100
120
140
160
0 2 4 6 8 10
Palatability (Pellets eaten)
Gro
wth
(%
mass i
ncre
ase y
ear
-1)
Fig. 9: Correlation between growth and defense.
CHAPTER 3:
PATTERNS OF RESOURCE ALLOCATION IN CARIBBEAN SPONGES: IS THERE
A TRADE-OFF BETWEEN REPRODUCTION AND DEFENSE?
40
ABSTRACT
On Caribbean coral reefs, some sponge species produce chemical defenses, while
others do not. Assuming resources are finite, species that produce defensive metabolites
would be expected to allocate fewer resources to growth and reproduction. In a previous
study, we documented a trade-off between growth and chemical defense among seven
branching sponges from shallow reefs off Key Largo, Florida. To investigate a trade-off
between reproduction and defense, we examined propagule output of seven species (six
branching and one vase-shaped) from November 2007 to October 2008. Each month,
tissue samples were collected from five individuals of the undefended species Iotrochota
birotulata, Niphates erecta, Callysponga armigera and Callyspongia vaginalis, and the
defended species Aplysina cauliformis, Aplysina fulva and Amphimedon compressa and
processed routinely for histology and light microscopy. For each sponge, a relative index
of reproductive output (ROI) was calculated as the percentage of reproductive propagules
out of the total tissue area scanned. Although reproductive output was highly variable, on
average, undefended and defended species had the same ROI. The lack of a trade-off
was attributed to the confounding factors, particularly the trade-off between propagule
formation and reproduction by fragmentation, that obscured the relationship between
propagule formation and defense. In combination with our previous studies of trade offs
between growth, reproduction and defense, we propose a conceptual model of how
resource allocation has influenced the evolution of sponge communities on Caribbean
coral reefs.
41
INTRODUCTION
Sponges are dominant members of the benthic sessile community on Caribbean
coral reefs (Targett and Schmahl 1984, Aronson et al. 2002, Maliao et al. 2008). Like all
living organisms, sponges allocate available resources to physiological functions such as
somatic growth and reproduction. In addition, some sponge species allocate resources to
produce secondary metabolites that deter predation (Paul 1992, Pawlik 1993). Yet, other
species that lack deterrent secondary metabolites co-exist on the coral reef despite
predation ((Pawlik et al. 1995, Pawlik 1998).
The variability in deterrent activity among sponge species can be likened to that
in terrestrial plants, where many hypotheses have been put forth to explain the
physiological and geographical patterns of chemical and physical defenses (Stamp 2003,
Agrawal 2007). According to the resource availability hypothesis, organisms allocate
available resources to defense, growth and reproduction (Coley et al. 1985, Bazzaz et al.
1987). When resources are limiting, a trade-off occurs. Resources invested in defense
must be diverted from growth or reproduction. Hence, organisms can either invest in
defense in order to resist predation, or tolerate predation by allocating resources to
somatic growth and the production of reproductive propagules. Such defense trade-offs
are well-documented in terrestrial plants (Koricheva 2002, Stamp 2003, Agrawal 2007)
For Caribbean coral reef sponges, the main predators are angelfishes, parrotfishes,
and turtles (Randall and Hartman 1968, Dunlap and Pawlik 1996, Leon and Bjorndal
2002), all of which feed on sponge species that lack chemical defenses. Predation is not
correlated with physical structures or nutritional quality in Caribbean reef sponges
(Chanas and Pawlik 1995), and with the exception of a few sponge species, the sharp
42
glass spicules found within sponge tissues generally do not serve a defensive function
(Burns and Ilan 2003, Jones et al. 2005). Secondary metabolites are the main agents
responsible for defense in sponges. Pawlik et al. (1995) tested the organic extracts of 73
Caribbean sponge species for palatability in feeding assays using the blue-head wrasse
(Thalassoma bifasciatum). From the results of lab and field assays, Caribbean reef
sponge species were grouped into three categories – preferred species are rapidly grazed
down and only survive in cryptic refugia, undefended and defended species both co-exist
on the reef (Pawlik 1998). The undefended species do not produce deterrent compounds,
whereas defended species produce a range of secondary metabolites to deter predation,
some of which have been isolated (Wright et al. 1991, Albrizio et al. 1995, Puyana et al.
2003, Nuñez et al. 2008).
Some resource trade-offs have been documented among Caribbean coral reef
sponges. Undefended sponge species grow faster (Leong and Pawlik, in prep) and have
faster rates of wound healing compared to defended species (Walters and Pawlik 2005).
After tissue damage, wound healing occurs to repair damaged tissue and prevent
microbial colonization and necrosis (Ayling 1983). Tissue repair occurs at a much faster
rate than somatic growth. Sponge species that are frequently grazed would be expected
to possess mechanisms for rapid wound healing in addition to faster rates of growth.
Sponge colonization patterns on new substrata also provide compelling evidence
for resource allocation trade-offs in Caribbean coral reef sponges. A survey was
conducted on the decks of a shipwreck four years after it was sunk to create an artificial
reef off Key Largo, Florida. Undefended sponge species dominated on the deck of the
wreck. The sponge community on the wreck differed from the nearest coral reefs, where
43
defended sponge species were present in high abundances along with undefended species
(Pawlik et al. 2008). A repeat survey 18 months later revealed that there were more
defended species beginning to recruit at low levels on the shipwreck. If undefended
sponge species produce more propagules or grow faster than defended species, then they
would recruit first and have a greater biomass than the defended sponge species,
consistent with the sponge community on the shipwreck.
Although a resource trade-off between growth and chemical defense has been
documented in Caribbean coral reef sponges (Leong and Pawlik, in prep), the trade-off
between reproduction and defense has not been directly measured. Hence, the aim of this
paper is to examine the resource trade-offs between reproduction and chemical defense
by comparing the propagule output of seven common species of Caribbean coral reef
sponges. The undefended species Iotrochota birotulata, Niphates erecta, Callysponga
armigera and Callyspongia vaginalis, and the defended species Aplysina cauliformis,
Aplysina fulva and Amphimedon compressa were used for this study. Apart from
Callyspongia vaginalis which grows as a cluster of tubes, all are branching sponges that
were previously used in the study of resource trade-offs between growth and chemical
defense. Of the defended sponge species, Aplysina spp. contain brominated tyrosine
derivatives similar to all members of the Verongidae (Puyana et al. 2003, Nuñez et al.
2008), and Amphimedon compressa produces a pyridinium alkaloid that is highly
deterrent to predators (Albrizio et al. 1995).
44
METHODS AND MATERIALS
Sponges were collected from Conch Wall in Key Largo, Florida, USA
(N24o56’44 W80
o27’23). Monthly samplings were carried out between November 2007
and October 2008 on the following dates: November 30, December 18, January 21,
February 24, March 28, April 19, June 1, June 27, August 1, August 28, September 26,
October 25. For each species, three 2cm x 2cm x 2cm blocks of tissue were cut from
each of five separate sponges and immediately fixed in 10% formalin buffered in
seawater. The samples were then routinely processed for histology. Dehydration was
carried out in gradated steps using ethanol (50%, 70%, 95%, 95%, 100%, 100%), and
samples were embedded in paraffin after passing through toluene as a clearing agent.
Sections were then cut with a rotary microtome at 10µm thickness, and stained using
haemotoxylin and eosin.
The slides were photographed using a SPOT digital camera connected to an
Olympus BX60 microscope at 4x magnification. A total area of 130mm2, corresponding
to 20 haphazard views among the sections, was photographed for each sponge. The area
of reproductive propagules (whether oocytes, embryos, or larvae) and the total area of the
slides were quantified using ImageJ imaging software (Rasband 1997). The
Reproductive Output Index (ROI = % area of propagules / total area of tissue scanned)
was calculated for each sample (after Whalan et al. 2007). The advantage of using ROI
over counts or other common measures of reproduction is that it enables comparisons of
reproductive output to be made between species.
Non-parametric statistical tests were used to determine whether total annual
reproductive output was different among the species, because ROI data contained a large
45
number of zeroes and did not have a normal distribution. The Kruskal-Wallis test was
used to analyze the data, which was followed by pair-wise comparisons using the
Wilcoxon rank-sum test with a Bonferroni correction.
RESULTS
Aplysina cauliformis and Aplysina fulva are oviparous, and produced small (12-
15µm) oocytes dispersed in the mesohyl. The remaining species are viviparous, and
samples containing oocytes, embryos and larvae were found. Callyspongia vaginalis,
Callyspongia armigera and Niphates erecta had propagules consolidated in brood
chambers, whereas Amphimedon compressa and Iotrochota birotulata had propagules
dispersed throughout the mesohyl. No sperm were observed in tissue sections from any
of the sponge species.
Among the seven sponge species, reproduction was highest between May and
October, peaking in July and August (Fig. 10). Amphimedon compressa contained
propagules throughout May to October. Callyspongia vaginalis also contained propagules
throughout the season, but had an additional reproductive peak in December. Niphates
erecta contained propagules in the early part of the season, between May to July.
Iotrochota birotulata contained propagules in the later half of the season, between July
and October. It was difficult to determine seasonality in Aplysina cauliformis, Aplysina
fulva and Callyspongia armigera because reproduction occurred at very low levels.
Aplysina cauliformis reproduced in June, but Aplysina fulva and Callyspongia armigera
were found to contain propagules in January, March and October which did not
correspond with the main reproductive season.
46
Annual reproduction varied widely between sponge species (Kruskal-Wallace;
χ2 = 53.244, df = 6, p<0.0001), and was highly variable within each species. The highest
levels of reproduction occur in Callyspongia vaginalis, the undefended tube sponge,
which had an annual ROI of 0.535 ± 1.082% (mean ± sd; n=60; Fig. 11). However, this
was not significantly different from the defended rope sponge Amphimedon compressa
(0.434 ± 1.166%) or the undefended rope sponge Iotrochota birotulata (0.391 ± 0.983%).
Significantly lower levels of reproduction were found in the undefended rope sponges
Niphates erecta (0.096 ± 0.427%) and Callyspongia armigera (0.041 ± 0.222%),
followed by the defended rope sponges Apysina cauliformis (0.001 ± 0.008%) and
Aplysina fulva (0.001 ± 0.010%).
The species with the highest ROI also had the highest number of reproductive
individuals (18 out of 60 for Callyspongia vaginalis, 15 out of 60 for Amphimedon
compressa, 17 out of 60 for Iotrochota birotulata, compared with 3 out of 60 for
Niphates erecta, 2 out of 60 for Callyspongia armigera and Aplysina cauliformis, and 1
out of 60 for Aplysina fulva). When calculated using only the sponges where propagules
were found, mean ROI per individual was comparable (0.012-0.019%) for all the
viviparous sponges, and much lower for the oviparous sponges (0.0003-0.0007%).
DISCUSSION
Reproductive outputs were consistent with other sponge species from the
literature. ROI for oocytes and larvae in the viviparous sponge species range from 0.01
to 0.535%, falling within the range of female ROI reported in the literature (Whalan et al.
2007). No sperm were observed in this study. The finding of little or no sperm has been
47
reported for other studies of reproduction in sponges, and is attributed to sampling bias
due to the transient nature of sperm in the mesohyl compared to the longer brooding
times of larvae (Fell 1989, Corriero et al. 1996, Tsurumi and Reiswig 1997). Whalan et
al. (2007) measured an ROI of 0.02-1.03% for female propagules in Rhopaloeides
odorabile, and calculated ROI of 1-12% for other sponge species in the literature. Low
reproduction for Aplysina spp. is in agreement with a study of Aplysina cauliformis by
Tsurumi and Reiswig (1997), in which only 10 of 208 sponges were found to contain
reproductive propagules.
Patterns of seasonality were also consistent with examples from the literature, in
which most sponges were observed to contain or release reproductive propagules
(oocytes, sperm or larvae) in the warmer months (Elvin 1976, Fromont 1994, Fromont
and Bergquist 1994, Mercurio et al. 2007, Whalan et al. 2007, McMurray et al. 2008). In
the present study, the reproductive season coincides with the season of highest growth
during the warmer months between May and October (Leong and Pawlik, in prep). Food
availability may be greater in the warmer months, enabling sponges to direct more
resources to both growth and reproduction.
The resource trade-off between reproduction and chemical defense among the
seven sponge species chosen for the present study is not as clear as that between somatic
growth and chemical defense in a previous study (Leong and Pawlik, in prep).
Comparing reproduction in seven Caribbean reef sponge species, defended sponge
species do not collectively produce less propagules than undefended species. If there is a
trade-off between reproduction and chemical defense, it is too weak to stand out from
among confounding factors such as concurrent trade-offs between reproduction and
48
growth among the sponge species. The trade-off between reproduction and defense is
well established in plants, but only when comparing among conspecifics (Bergelson and
Purrington 1996, Koricheva 2002).
The oviparous sponges Aplysina spp. do not allocate much resources to
reproduction, with only a few individuals producing a small number of tiny oocytes.
Lower resource allocation to reproduction leaves resources to be diverted to other
functions such as growth and chemical defense. Aplysina spp. are highly defended
sponges with very high growth rates (Leong and Pawlik, in prep). Based on phylogenetic
studies, oviparity has arisen multiple times in the Demospongiae (Borchiellini et al. 2004).
Oviparous species broadcast spawn oocytes and sperm into the water column, and no
further investment goes into larval development. Viviparous species brood larvae, which
can grow up to 0.5mm in size before release (Leys and Ereskovsky 2006).
Among the viviparous sponges, the trade-off between reproduction and chemical
defense may be obscured by other trade-offs involving reproduction. Organisms that
reproduce by asexual fragmentation allocate fewer resources to propagule formation
(Tunnicliffe 1981, Highsmith 1982, Lasker 1984, Thomsen and Hakansson 1995, Barrat-
Segretain et al. 1998). Branching sponge species can disperse and rapidly colonize new
substratum by fragmentation, and therefore allocate more resources to vegetative growth,
producing fewer propagules (Wulff 1991, Wulff 1995, Tsurumi and Reiswig 1997, Leong
and Pawlik, in prep). The undefended sponge Callyspongia armigera has one of the
lowest ROI, but the highest growth rate (Leong and Pawlik, in prep). Its congener,
Callyspongia vaginalis, has high ROI but lower growth rates (Leong and Pawlik, in prep).
Trade-offs in reproduction between propagule formation and fragmentation would
49
confound the relationship between chemical defenses and propagule formation in
branching sponge species, of which all but one were used in the present study.
In order to evaluate resource allocation patterns between chemical defense,
somatic growth and reproduction, respective values of each of the seven sponge species
were plotted on a 3D graph with the three factors on the axes (Fig. 12). Values for
chemical defense were taken from Pawlik et al. (1995). Pawlik et al. (1995) used a
palatability index indicating the mean number of food pellets eaten in aquarium assays
using a generalist reef fish (Thalassoma bifasciatum). To obtain a corresponding index
of defense for each sponge species, the mean number of pellets rejected was calculated
from the palatability index by subtracting the mean number of pellets eaten from the total
number of pellets. For each axis, the maximum value recorded was set at a value of 90,
and the remaining values were scaled between 0-90 (i.e. a defense of 10 is 90, 8 is 72, 5
is 45, and so forth). A theoretical surface plot where all axes sum to 100 was overlain on
Fig. 12 in the form of a net (i.e. the points 90, 10, 0; 50, 50, 0 and 33, 33, 33 all lie on the
surface plot, which is shaped like a triangle with the points representing exclusive
allocation to one of the three factors growth, reproduction or defense). This surface
represents the range of values to which sponge species are expected to allocate their
resources. Most species should lie on this theoretical plane since to lie below it would be
uncompetitive, and species are constrained from lying above the plane by the total
amount of resources available. Species that lie above the plane should be more effective
survivors and more abundant than species that lie on or below it.
Sponge species disperse widely across the chart (Fig. 12), indicating that sponges
employ a range of evolutionary strategies to cope with resource allocation constraints.
50
For example, sponges species such as Aplysina cauliformis and Aplysina fulva have high
defense, but low growth and reproduction and lie on the top of the chart. Conversely, the
undefended species Callyspongia vaginalis and Callyspongia armigera lie near the
bottom. The two are at the ends of the triangle formed by the theoretical surface plot,
which represents either higher allocation to growth (near corner) or reproduction (far
corner). The small number of species on the chart limits our ability to observe any other
patterns in the spread of points. Defended sponges have the highest potential to move
above the plane, and defense probably evolved separately in different groups of sponges
(Wright et al. 1991, Albrizio et al. 1995, Puyana et al. 2003, Nuñez et al. 2008) for this
reason. There are several assumptions of the model that need to be investigated further.
The model does not account for variation in filtration and resource uptake rates among
the different species, which may explain why defense, reproduction and growth do not
sum to 100 in all species. Secondary metabolites used for chemical defense are expected
to be costly to produce due to the requirement for raw materials, the production and
storage of metabolites, and the prevention of autotoxicity (Van Alstyne et al. 2001). Yet,
the exact cost of chemical defense is difficult to work out. Resource allocation (e.g.
energy or carbon invested in defense) should be constrained by natural selection to be
correlated with phenotypic traits (e.g. palatability), but the maximum levels of defense
may not cost much resources to produce compared with growth and reproduction, in
which case the axes for Fig. 12 should be scaled differently. Aplysina spp. and
Amphimedon compressa, both defended species, had high comparative growth rates and
reproductive outputs respectively.
51
Our current understanding of trade-offs in resource allocation to defense, growth
and reproduction can be summarized in a conceptual model (Fig. 13). Clear trade-offs
between growth and defense, and between reproduction and growth have been
demonstrated among Caribbean sponge species. The trade-off between reproduction and
defense is less clear. With this new information in hand, we can revisit and attempt to
explain observations of sponge colonization on a shipwreck off Key Largo, Florida
(Pawlik et al. 2008). On the shipwreck, sponge species with the highest abundance
correspond with the most prolific undefended sponge species, the branching sponge
Iotrochota birotulata and the tube sponge Callyspongia vaginalis. In addition to
recording sponge cover on the shipwreck, Pawlik et al. (2008) also measured the volumes
of the largest sponges of each species present on the shipwreck. The largest sponges
present were the tube sponge Callyspongia vaginalis and its congener Callyspongia
fallax. Although Iotrochota birotulata was more abundant on the ship, individuals were
smaller in size. In the branching species, fragmentation of the larger individuals would
result in smaller individuals. Considering that the other branching species studied have
lower rates of propagule formation, it is possible that recruitment occurred later for these
species, explaining their lower abundances. Sponge fragments, which are much larger
than propagules, are unlikely to travel to the shipwreck from the nearest reef 800m away.
Fragments are even more unlikely to make it onto the deck, which sits 15m above the sea
floor. It is expected that branching species, once recruited to the shipwreck, would
rapidly increase their abundance by growing and fragmenting. One question that arises is
why Amphimedon compressa, which produces many propagules and is common on
nearby reefs, was not more abundant on the shipwreck. Information about larval
52
dispersal distances and settlement behavior could help to answer this question. With the
exception of Amphimedon compressa, the sponge community on the shipwreck is well
explained by resource allocation trade-offs. The low abundance and late recruitment of
defended sponge species on the shipwreck strongly suggests that there is a trade-off
between reproduction and chemical defense in Caribbean coral reef sponges, and we
were unable to detect it because of confounding factors due to the morphology of the
sponge. The reproduction-defense trade-off would possibly be more detectable by
comparing non-branching sponge species. It will be interesting to see how the sponge
community continues to change on the shipwreck, and whether future sponge community
patterns will corroborate our conceptual model of resource allocation trade-offs.
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56
0
0.5
1
1.5
2
2.5
Nov-0
7
Dec-0
7
Jan-0
8
Feb-0
8
Mar-
08
Apr-
08
May-0
8
Jun-0
8
Jul-08
Aug-0
8
Sep-0
8
Oct-
08
Mean
RO
I
Callyspongia armigeraCallyspongia vaginalisIotrochota birotulataNiphates erectaAmphimedon compressaAplysina cauliformisAplysina fulva
Fig. 10: Mean monthly ROI for seven coral reef sponge species in Key Largo, Florida.
n=5
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
1.8
Cally
spongia
arm
igera
Cally
spongia
vagin
alis
Iotr
ochota
birotu
lata
Nip
hate
s
ere
cta
Am
phim
edon
com
pre
ssa
Aply
sin
a
caulif
orm
is
Aply
sin
a
fulv
a
Mean
RO
I
BB BA BAA
Fig. 11: Total yearly ROI for seven coral reef sponge species in Key Largo, Florida.
n=60, means + sd. Post-hoc comparisons were carried out using Wilcoxon’s test with a
Bon-ferroni correction. Different letter groups indicate a statistical difference was found.
Bold letters indicate defended species.
57
Fig. 12: Plot of growth, reproduction and defense. For each axis, the maximum value
occurring was set at 90 and all other values were scaled between 0-100. Defended
species (dots): Aplysina cauliformis, Aplysina fulva (solid dots), Amphimedon compressa
(hollow dot). Undefended sponges: Callyspongia armigera (solid triangle), Callyspongia
vaginalis (hollow triangle), Iotrochota birotulata (solid square), Niphates erecta (hollow
square). Theoretical surface plot is overlaid, where the three axes sum to 100 (black
mesh).
Fig. 13: Conceptual model of trade-offs between defense, growth and reproduction in
sponges.
Reproduction
Growth
Defense Morphology
Tolerance/Resistance
Tolerance/ Resistance