Filter Feeding Ecology of Erect Branching Sponges on Caribbean Coral Reefs Anna Margaret Strimaitis

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2012 Filter Feeding Ecology of Erect Branching Sponges on Caribbean Coral Reefs Anna Margaret Strimaitis

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COLLEGE OF ARTS AND SCIENCE

FILTER FEEDING ECOLOGY OF ERECT BRANCHING SPONGES ON CARIBBEAN

CORAL REEFS

ANNA MARGARET STRIMAITIS

A Thesis submitted to the Department of Biological Science in partial fulfillment of the requirements for the degree of Master of Science

Degree Awarded: Summer Semester, 2012

Anna Strimaitis defended this thesis on April 25, 2012.

The members of the supervisory committee were:

Janie L. Wulff Professor Directing Thesis

Markus Huettel University Representative

Don R. Levitan Committee Member

Alice A. Winn Committee Member

Kay M. Jones Committee Member

The Graduate School has verified and approved the above-named committee members, and certifies that the thesis has been approved in accordance with university requirements.

For Aunt Judy. (November 21, 1945 – September 8, 2008)

Your fondness for the sea and the Seminoles is always in my thoughts.

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ACKNOWLEDGEMENTS

I thank Janie Wulff for her collaboration in the design and execution of this project. I thank Markus Huettel, Kay Jones, Don Levitan, and Alice Winn for discussion on project development, data analysis, and writing. Special thanks to Ruth Didier for her guidance and expertise with the flow cytometry analysis at the Florida State University College of Medicine Flow Cytometry Facility. I also thank Cedric Magen for analyzing the DOC and TN samples. From Florida State University, I thank Sonja Bridges and FSU ADP, Greg Hoffman, Audrey Nichols, Cheryl Pye, and Roy Weidner for their help in designing, building, ordering, and preparing field research equipment and facilitating use of research gear. Special thanks to Judy Bowers for everything that she does for the Biological Science graduate students. From the Caribbean Coral Reef Ecosystems (CCRE) Program at Carrie Bow Cay, I thank Craig Sherwood, Zachary Foltz, and Michael Jones, and from The Smithsonian Tropical Research Institute at Bocas del Toro, I thank Carlo Avila, Plinio Gondola, Urania Gonzalez, Javier Jara, Gabriel Jacome, Michael Lang, Juan Mate, and Edgardo Ochoa. Both groups provided excellent logistical support necessary to carry out these experiments and keep samples frozen at remote field stations. I also thank the Curacao Sea Aquarium and CARMABI for support in Curacao. Special thanks to Brendan Biggs for hours of field assistance in Curacao and Bocas del Toro and to Colin Wulff for field assistance in Belize. Kacey Grace completed the AFDW analysis for D. anchorata. Nicole Poulton at the Bigelow Laboratory for Ocean Sciences advised on water sample collection and analysis with flow cytometry. This research was funded by two Loftin Foundation Awards from Florida State University Department of Biological Science to Anna Strimaitis and the National Science Foundation under Grant No. 0550599 to Janie Wulff. Special thanks to Brendan Biggs and Tim Swain for their never-ending support, advice, and encouragement as fellow lab mates and friends. A big thank you also to the enthusiastic academic and social support from the biology graduate student community past and present, particularly EERDG, Mia Adreani, Denis Avey, Lindsey Biggs, Emily Field, Nicole Fogarty, Bonnie Garcia, Elise Gornish, Josh Bear Grinath, Lisa Hollensead, Katie Lotterhos, Megan Lowenberg, Christina Kwapich, Ariel Simonton, Jennifer Schellinger, and, last but certainly not least, Caroline Stahala. Finally, thank you to my family for their unconditional encouragement, for their sincere interest in my research, and for raising me as an inquisitive scientist.

TABLE OF CONTENTS

List of Tables ...... vi List of Figures ...... viii Abstract ...... ix INTRODUCTION ...... 1 1. PICOPLANKTON ABUNDANCE IN WATERS AVAILABLE TO FILTER FEEDING COMMUNITIES OF CARIBBEAN CORAL REEF, MANGROVE, AND SEA GRASS ECOSYSTEMS...... 3 1.2 Methods...... 5 1.3 Results ...... 11 1.4 Discussion ...... 25 2. FILTER FEEDING RATES AND SELECTIVITY OF ERECT BRANCHING SPONGES ON CARIBBEAN CORAL REEFS ...... 29 2.2 Methods...... 35 2.3 Results ...... 40 2.4 Discussion ...... 47 CONCLUSION ...... 52 REFERENCES ...... 53 BIOGRAPHICAL SKETCH ...... 61

LIST OF TABLES

1.1 Ambient water sample counts and days of collection ...... 9

1.2 Panama and Belize summer light intensity ANOVA table ...... 11

1.3 Belize December and May light intensity ANOVA table ...... 12

1.4 Dissolved nitrogen ANOVA table ...... 13

1.5 Total picoplankton cell density ANOVA table ...... 14

1.6 Picoplankton prey type cell density ANOVA table ...... 14

1.7 Picoplankton trophic group cell density ANOVA table ...... 14

1.8 Total picoplankton carbon ANOVA table ...... 15

1.9 Picoplankton prey type carbon ANOVA table ...... 16

1.10 Picoplankton trophic group carbon ANOVA table ...... 16

1.11 Total picoplankton nitrogen ANOVA table ...... 17

1.12 Picoplankton prey type nitrogen ANOVA table ...... 17

1.13 Picoplankton trophic group nitrogen ANOVA table ...... 17

2.1 Size, carbon, and nitrogen content of different picoplankton prey types ...... 32

2.2 Significance of grazing ANOVA table ...... 40

2.3 Pairwise comparisons of prey growth in control chamber to each treatment chamber ...... 40

2.4 MANOVA table comparing observed sponges diet to expected diet based on ambient water ...... 42

2.5 ANOVA table for carbon in the sponge diet ...... 42

2.6 ANOVA table for nitrogen in the sponge diet ...... 43

2.7 ANOVA table for carbon in the Desmapsamma anchorata diet ...... 43

2.8 ANOVA table for nitrogen in the Desmapsamma anchorata diet ...... 43

2.9 ANOVA table for carbon in the Amphimedon compressa diet ...... 44

2.10 ANOVA table for nitrogen in the Amphimedon compressa diet ...... 44

2.11 ANCOVA table for cyanobacteria clearance rate ...... 46

2.12 ANCOVA table for heterotroph clearance rate ...... 46

2.13 ANCOVA table for prochlorophyte clearance rate ...... 46

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LIST OF FIGURES

1.1 Map of the Caribbean depicting the relative locations (*) of the three main study locations: Belize, Panama, and Curacao ...... 5

1.2 Map of Carrie Bow Cay, 10-15 km from mainland Belize ...... 6

1.3 Map of Bocas del Toro, Panama ...... 7

1.4 Map of Curacao ...... 7

1.5 (a) Mean summer light intensity in Panama and Belize and (b) mean light intensity in winter and summer in Belize ...... 12

1.6 (a) Mean DOC and (b) mean TN concentrations at the mangrove and reef sites in Panama and Belize...... 13

1.7 Mean (± 1 s.e.) picoplankton cell density at each site, partitioned by prey type...... 15

1.8 Mean (± 1 s.e.) carbon content of picoplankton at each site, partitioned by prey type ...... 16

1.9 Mean (± 1 s.e.) nitrogen content of picoplankton at each site, partitioned by prey type ...... 18

1.10 (a) Mean picoplankton cell density at each Panama site, partitioned by prey type ...... 19

1.11 (a) Mean picoplankton cell density at each Belize site, partitioned by prey type ...... 21

1.12 (a) Mean picoplankton cell density at each reef site, partitioned by prey type ...... 23

1.13 (a) Mean picoplankton cell density at each mangrove site, partitioned by prey type ...... 24

2.1 Images of the internal canal architecture ...... 32

2.2 Study sites in (a) Belize and (b) Panama ...... 35

2.3 (a) Sponge fragments healing on PVC stands and (b) experimental chambers designed to quantify sponge filter feeding rates and selectivity ...... 37

2.4 Mean growth for each prey type in the control chamber and chambers with a sponge species present ...... 41

2.5 Comparison between the observed carbon and nitrogen contributed by each prey type to the sponge diet and the expected contribution based on the composition of the ambient water ...... 45

2.6 Relationships between sponge cyanobacteria clearance rate and sponge size, heterotroph clearance rate and sponge size, and prochlorophyte clearance rate and sponge size ...... 47

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ABSTRACT Sponges are unique filter feeding organisms with complex canal and flagellated chamber aquiferous systems. These systems allow them to specialize in clearing the smallest plankton size class (picoplankton) from the water. Sponges serve many important ecological functions, but they may be best known for efficiently filtering picoplankton, such as phytoplankton bloom species, from the water column. This ecological function increases water clarity and allows more light to penetrate to photosynthesizing corals and sea grasses. As abundant and efficient members of the benthic filter-feeding community, it is important to understand how and to what extent sponges can maintain water quality on healthy coral reefs and restore water quality on declining coral reefs. The first goal of this research was to quantify and compare the abundance of picoplankton resources, in terms of number of cells and carbon and nitrogen content, available to sponge filter feeders in Caribbean coral reef, mangrove, and sea grass habitats. The four primary picoplankton prey types (picoeukaryotes, cyanobacteria, prochlorophytes, and heterotrophic bacteria) were quantified using flow cytometry. Data were also collected on light intensity and dissolved nutrient concentrations because some sponge species harbor phototrophic and heterotrophic bacterial symbionts that can utilize these resources. We studied sites in all three habitats near Carrie Bow Cay, Belize, two reef sites and one mangrove site in Bocas del Toro, Panama, and one reef site in Curacao. These sites were chosen specifically to explain observed patterns in long-term sponge growth rate data that suggest resource limitation. Overall, heterotroph cell densities and carbon and nitrogen were greater at the mangrove sites than at the reef sites, but autotroph cell densities and carbon and nitrogen at the reefs sites were greater than or equal to those at the mangrove sites. Furthermore, there were greater heterotroph and autotroph cell densities and carbon and nitrogen at the reef sites in Bocas del Toro than at the reef sites in Belize and Curacao, and the autotroph cell densities and carbon and nitrogen were also greater at the mangrove site in Bocas del Toro than the mangrove site in Belize. The second goal of this research was to quantify and compare the effect of sponge grazing on the four picoplankton prey types, the clearance rate of each prey type for each species, and selectivity (i.e. clearance rate standardized by relative abundance of prey types in the ambient water). These variables were quantified and compared for six of the most common erect branching sponge species representing three different orders at the reef sites in Belize and

Panama: Aplysina cauliformis (Order Verongida), Aplysina fulva (Order Verongida), Desmapsamma anchorata (Order Poecilosclerida), Iotrochota birotulata (Order Poecilosclerida), Niphates erecta (Order Haplosclerida), and Amphimedon compressa (Order Haplosclerida). Sponge feeding was quantified using chamber experiments that compared the change in prey density in the course of 20 minutes in a chamber with a filtering sponge to the change in prey density in the course of 20 minutes in a control chamber without a sponge. All six sponge species significantly removed cyanobacteria and heterotrophic bacteria compared to controls, none of the species removed picoeukaryotes, and all species except the two Aplysina species removed prochlorophytes. The two Aplysina species also cleared cyanobacteria more slowly than all other sponge species. Amphimedon compressa and Niphates erecta cleared heterotrophs more quickly than the other species and Amphimedon compressa also cleared cyanobacteria more quickly than all other sponge species. Finally, Aplysina fulva and Aplysina cauliformis removed heterotrophic bacteria more efficiently than other prey types, Iotrochota birotulata and Desmapsamma anchorata removed cyanobacteria more efficiently than other prey types, and Amphimedon compressa and Niphates erecta did not demonstrate selectivity for any of the prey types. This is the first study to quantify picoplankton abundances in the water directly available to sponges in reef, sea grass, and mangrove habitats and to also quantify the feeding ecology of six erect branching sponge species at multiple reef sites in the Caribbean. The differences documented in the picoplankton communities between sites indicate that differences in growth rates for individuals of the same species between sites may be controlled by bottom-up forces. Furthermore, if sponges partition picoplankton resources as this study suggests, then diverse sponge communities are essential to maintain water quality in these systems by effectively removing all types of picoplankton. Finally, sponges fulfill important ecological functions in these ecosystems, and it is possible that their filtering power could be harnessed in a species- specific manner for remediation efforts to improve water quality in these ecosystems.

INTRODUCTION

Marine plankton represents an abundant and available food source that only certain animals can efficiently extract. Filter feeding organisms have evolved structures or morphologies well adapted to utilize resources suspended in the water column (Gili and Coma 1998). This trophic interaction represents an important energy transfer process in coastal marine ecosystems (Koseff et al. 1993) as dilute concentrations of plankton biomass in the pelagic food web are converted into animal biomass in the benthic food web. The picoplankton size class (plankton < 2 μm) can dominate the entire planktonic community in terms of both biomass and production (Stockner and Antia 1986). In addition to their abundance, the composition and trophic interactions of the picoplankton group are important to understand because this group includes microorganisms of ecological concern, like species of phytoplankton blooms that are fueled by coastal nitrogen and phosphorous runoff, as well as pathogenic microbes, likes bacteria and viruses (Sherr & Sherr 1991, Peterson et al. 2006, Maldonado et al. 2010). Sponges (Phylum Porifera) are one of the dominant filter feeding invertebrate groups on Caribbean coral reefs, where sponge volume exceeds that of corals (Wulff 1994). Sponges are unique because they specialize in retaining picoplankton as small as 0.1 μm with nearly 100% efficiency (Reiswig 1971, 1975). While it is well established that many benthic suspension feeders, like bivalves, are selective in their grazing (Cranford and Gordon 1992, Ward and Shumway 2004), there is a persisting notion that sponges must be nonselective filter feeders. This idea stems from the presumption that the microvilli of the choanocytes act as a true sieve, with no mechanism for particle selection, yet there is some preliminary evidence of selective particle retention in certain sponge species (Reiswig 1971, Ribes et al. 1999). Given the ecological importance of the trophic interaction between picoplankton and sponges, this research consisted of two related projects. Chapter 1 focused on quantifying the picoplankton resources available, in terms of both cell density and carbon and nitrogen equivalents, to sponges in Caribbean coral reef, mangrove, and sea grass ecosystems in Panama, Belize, or Curacao. The questions addressed were: Do mangrove sites have more picoplankton resources than reef sites? Do sites in Bocas del Toro, Panama have more resources than sites in Belize and Curacao? Are the relative abundances, in terms of number of cells and carbon and nitrogen equivalents of the different picoplankton prey types similar across sites? Chapter 2

focuses on the chamber experiments that were conducted to quantify the filter feeding rates and selectivity of multiple sponge species at coral reef sites in Panama and Belize. The questions addressed were: Do these sponge species effectively graze all four picoplankton prey types? Do these sponge species filter selectively? Do these sponge species remove prey at similar rates? These basic aspects of sponge filter feeding ecology are currently unknown. If this knowledge is paired with information on picoplankton abundances at different Caribbean coral reef sites, we can begin to address broader, ecosystem level questions such as: What is the effect on the water column of these species at documented densities on Caribbean reefs? How are the distribution and abundance of these sponge species across Caribbean reef sites controlled by the composition of the water column? How can we use species specific information on clearance rates and selectivity to inform bioremediation efforts on declining coral reefs?

CHAPTER ONE

PICOPLANKTON ABUNDANCE IN WATERS AVAILABLE TO FILTER FEEDING COMMUNITIES OF CARIBBEAN CORAL REEF, MANGROVE, AND SEA GRASS ECOSYSTEMS

Introduction

Planktonic bacteria and other microorganisms play important ecological roles in the cycling of carbon and nitrogen within marine environments (Sorokin 1981, Azam et al. 1983, Cole et al. 1988, Sherr and Sherr 1991). It is necessary to know about the composition of the planktonic community in different environments in order to understand food webs and biogeochemical fluxes (Sherr and Sherr 1991). Marine plankton represents an abundant and available food source that only certain animals can efficiently extract. Filter feeding organisms have evolved structures or morphologies well adapted to utilize resources suspended in the water column (Gili and Coma 1998). This trophic interaction represents an important energy transfer process in coastal marine ecosystems (Koseff et al. 1993) as dilute concentrations of plankton biomass in the pelagic food web are converted into animal biomass in the benthic food web. Studies estimating in situ particle uptake by marine communities indicate that plankton grazing is a very important component of benthic-pelagic coupling in marine environments from the Red Sea to Hawaii to Australia to the Caribbean (Gast et al. 1998, Yahel et al. 1998, Ribes et al. 2003, Lesser 2006, Patten et al. 2011, Strimaitis and Wulff, in prep). The picoplankton size class (plankton < 2 μm) is one of two size classes that dominate the entire planktonic community by as much as 60-80% of the biomass and 44-90% of the production in Atlantic and Pacific tropical oceans (reviewed in Stockner and Antia 1986). This importance has motivated research studying picoplankton dynamics and interactions with other plankton size classes (Sherr and Sherr 1991, Van Wambeke et al 1996). In addition to their abundance, the composition and trophic interactions of the picoplankton group are important to understand because this group includes pathogenic microbes, like bacteria and viruses, as well as microorganisms of ecological concern, like species of phytoplankton blooms that are fueled by coastal nitrogen and phosphorous runoff (Sherr & Sherr 1991, Peterson et al. 2006, Maldonado et al. 2010).

The significance of picoplankton as a nutrient resource for filter feeding animals of marine benthic communities (Reiswig 1971, Wilkinson et al. 1984, Ribes et al. 1999, Peterson et al. 2006, Lesser 2006, Yahel et al. 2006) highlights the importance for quantifying picoplankton concentrations available to filter feeding animals at sites with different physical properties, chemical characteristics, and coastal or anthropogenic influences. Sponges (Phylum Porifera) are likely the most abundant filter feeders in biomass (Wulff 1994) that are able to specialize on efficiently removing particles in the picoplankton size range (Reiswig 1971, 1975, 1990, Ribes et al. 1999). Studies demonstrating that some tube shaped sponges exhibit greater growth rates at increasing depth where there is a greater food supply (Lesser 2006, Trussell et al. 2006) provide some evidence that sponges may be resource limited. Furthermore, other studies following growth rates of the many erect branching sponge species found on reefs at Bocas del Toro, Panama and reefs at Blue Ground Range, Belize have found that individuals of the same sponge species grow faster in Panama than in Belize (Wulff, in prep.). Growth rates for individuals of the same species were also faster when fragments were transplanted from reef to mangrove habitats (Wulff, in prep). These studies raise questions of whether the distribution and abundance of sponge species is influenced by bottom up forces and whether there is more picoplankton available to filter feeding sponges in Panama than other Caribbean reefs and in mangrove habitats compared to reef habitats at these locations. In this study, we analyzed the composition and abundance (in terms of individual cells and carbon and nitrogen content of cells) of the picoplankton community in the water ambient to filter feeders at sites in different habitats (coral reef, mangrove, sea grass) and locations (Carrie Bow Cay, Belize; Bocas del Toro, Panama; and Curacao; Fig 1.1) at which sponge growth rates have been quantified and compared. Based on this growth rate information, we predicted that picoplankton would be more abundant at these mangrove sites than at these reef and sea grass sites, and that there would be more picoplankton at the Bocas del Toro, Panama sites than at the Belize and Curacao sites. We had the opportunity to sample in both winter and summer at the Belize sites and took advantage of this chance to collect more data that may inform how picoplankton abundances vary seasonally over tropical marine habitats. Other factors such as light intensity and dissolved nutrient concentrations may influence the picoplankton resources available to filter feeding sponges, because the autotrophic prey types photosynthesize and picoplankton can utilize dissolved nutrients. Solar radiation has been previously shown to inhibit

heterotrophic bacterial production to a depth of about 5 m (Visser et al. 1999). Furthermore, some sponges are termed “bacteriosponges” because they harbor dense symbiotic populations of both phototrophic and heterotrophic microbes that may transfer nutrition to their sponge hosts (Wilkinson and Garrone 1980, Reiswig 1981, Wilkinson 1983, Diaz & Ward 1997, Yahel et al. 2003, Hoffman et al. 2005, de Goeij et al. 2008, Freeman and Thacker 2011). We predict that sites with more intense light will have more autotrophic picoplankton and sites with more dissolved nutrients will have more heterotrophic picoplankton.

Figure 1.1. Map of the Caribbean depicting the relative locations (*) of the three main study locations: Belize, Panama, and Curacao. Figure adapted from Google Maps.

Methods

Study Sites The study in Belize included a coral reef site at 3 m depth (Blue Ground Range; site BZE-R), a mangrove site at 0.8 m depth (Twin Cays; site BZE-M), and a sea grass site at 2 m depth (mouth of Twin Cays; site BZE-SG). All sites were accessed from the Smithsonian field station on Carrie Bow Cay, 20 km from mainland Belize (Fig 1.2). The study in Panama included two coral reef sites and a mangrove site located in a shallow lagoon system directly off

the coast of the Smithsonian Tropical Research Institute Bocas del Toro field station, which receives high nutrient runoff from the nearby mountains and developing coastline towns. One reef site was at a depth of 3 m (site PAN-R1) located across from the field station and about 10 m in distance from the mangrove site at 0.8 m depth (site PAN-M). Both of these sites are located at the opening of the bay leading to the relatively densely populated town of Bocas del Toro. The second reef site in Bocas del Toro was at a depth of 6 m located near STRI point (site PAN-R2), which is about 2 km from the town of Bocas del Toro (Fig 1.3). While the study sites in Bocas del Toro are referred to here as the Panama sites, we emphasize that the picoplankton abundance found at Bocas del Toro is likely not representative of all shallow Caribbean reefs in Panama because of Bocas del Toro’s unique natural and anthropogenic influences. The study in Curacao was conducted on one coral reef at a depth of 6 m (site CUR-R) off shore from the Curacao Sea Aquarium (Fig 1.4). These sites were chosen specifically to inform observed patterns in long term sponge growth rate data that suggest resource limitation.

Figure 1.2. Map of Carrie Bow Cay, 10-15 km from mainland Belize. Study sites (BZE-R, BZE-M, BZE-SG) are indicated. Figure adapted from Smithsonian Institution CCRE Annual Report 2011.

Figure 1.3. Map of Bocas del Toro, Panama. Study sites (PAN-R1, PAN-R2, PAN-M) are indicated. The developing town of Bocas del Toro is indicated by the lined region. Figure adapted from Google Maps.

Figure 1.4. Map of Curacao. Reef study site at Curacao Sea Aquarium is indicated. Figure adapted from Van Duyl & Gast 2001.

Light Intensity Light intensity (lux) measurements were recorded at each site in Belize and Panama using HOBO data loggers. Data were collected in Belize for twelve days in December of 2009 and for five days in May of 2010. Data were collected in Panama for nine days in August of 2011.

Water Sample Collection and Analysis Between 3 and 30 replicate water samples were collected from all sites in Belize, Panama, and Curacao on multiple days to quantify the picoplankton composition of the water ambient to filter feeding communities at the reef and sea grass sites and amidst the filter feeding communities on prop roots at the mangrove sites (Table 1.1). When samples were collected to quantify the picoplankton abundances in the ambient water, three replicate samples were taken. We also conducted sponge feeding experiments, as described in Chapter 2, and during those experiments, it was necessary to collect a zero time point sample for every experiment. That zero time point also quantified picoplankton in the ambient water column. Because of these zero time point samples, the days when we ran feeding experiments have much larger sample sizes (Table 1.1). At the Belize mangrove and reef sites (BZE-M and BZE-R), ambient water samples were collected on two days in December 2009 and feeding experiments were conducted on 1 day in May 2010. At the Belize sea grass site (BZE-SG), ambient water samples were collected on 1 day in both December 2009 and May 2010. At the Panama mangrove and nearby reef site (PAN-M and PAN-R1), feeding experiments were conducted on 1 day in August 2010 and ambient water samples were collected on 3 days in August 2011. At the STRI point reef site (PAN-R2), feeding experiments were conducted on 1 day and ambient water samples were collected on 2 other days in August 2011. Finally, in Curacao ambient water samples were collected on 1 day and feeding experiments were conducted on another day in July 2011. The feeding experiments conducted at the reef sites in Belize and Panama are described in Chapter 2.

Table 1.1 Ambient water sample counts and days of collection Site Day 1 Day 2 Day 3 Day 4 Total BZE-M Dec 2009 (N=3) Dec 2009 (N=3) May 2010 (N=30) NA N=36 BZE-R Dec 2009 (N=3) Dec 2009 (N=3) May 2010 (N=20) NA N=26 BZE-SG Dec 2009 (N=3) May 2010 (N=3) NA NA N=6 PAN-M Aug 2010 (N=24) Aug 2011 (N=3) Aug 2011 (N=3) Aug 2011 (N=3) N=33 PAN-R1 Aug 2010 (N=24) Aug 2011 (N=3) Aug 2011 (N=3) Aug 2011 (N=3) N=33 PAN-R2 Aug 2011 (N=3) Aug 2011 (N=3) Aug 2011 (N=30) NA N=36 CUR-R July 2011 (N=3) July 2011 (N=19) NA NA N=22

A 25 mL syringe was used to collect water from each site. After collection, each syringe was mixed by inversion before 1.7 mL was filtered through a 100-μm mesh into a 2-mL cryovial and fixed with formaldehyde to a final concentration of 0.5%. Samples in cryovials were allowed to fix on ice in a UV-proof box for at least one hour and no more than eight hours. After fixation, samples were flash frozen in liquid nitrogen at the field station, transported back to Florida State University on dry ice or in a dry shipper, and stored at -80οC. For dissolved organic carbon (DOC) and total nitrogen analysis (TN), 15 mL of the syringe sample was filtered through a pre-combusted GF/F glass fiber filter. The filtered water was preserved with HCl to a pH of 2 and stored in a dark refrigerator until analysis at Florida State University with a Shumadzu TOC-V analyzer with ASI autosampler. DOC and TN samples were only collected during August 2010 sampling at Panama reef and mangrove sites (PAN-R1 and PAN-M), and during May 2010 sampling at Belize reef and mangrove sites (BZE-R and BZE-M). In collaboration with The Florida State University College of Medicine, flow cytometry was used to quantify heterotrophic bacteria, Synechococcus-like cyanobacteria, Prochlorococcus-like prochlorophytes, and picoeukaryote cells. Water samples were analyzed using a FACSCanto flow cytometer equipped with two lasers and six colors. SYBR Green I (a nucleic acid gel stain) was used to stain DNA according to Crain (2003). Five parameters were collected in list mode on a log scale: red fluorescence with a Blue 670LP laser (from chlorophyll a), orange fluorescence with a Blue 585/42 laser (from phycoerythrin), green fluorescence with a FITC laser (from DNA stained with SYBR Green I), forward and side angle light scatter signals (influenced by particle size and shape, refractive index, and internal cell structure). To analyze

the autotrophic fraction of the picoplankton (cyanobacteria, prochlorophytes, and picoeukaryotes), the water sample was run one time for five minutes at high flow rate triggered by the Blue 670LP laser. To analyze all DNA containing particles, the sample was run one time for three minutes on low flow rate triggered by the FITC laser. Flow rates were chosen to maintain particle counts less than 1000 counts per second to reduce error. The cell counts for all quantified picoplankton populations were converted to density using the known volume of water that was analyzed by the flow cytometer in the controlled amount of time. The density of heterotrophic cells was calculated by subtracting the density of phototrophic cells in that sample from the total density of cells.

Statistical Analysis All statistical analyses were conducted using R statistical software (R Developmental Core Team 2011). All data were tested for assumptions of normality and homogeneity of variance and, if they did not meet the assumptions, data were transformed when possible or nonparametric methods were used. A 1-way ANOVA was used followed by Tukey Post Hoc tests to compare average light intensity (lux) measurements for each Panama site and each Belize site sampled in May. A separate analysis was run to compare May and December in Belize. Average light intensity was log transformed to meet assumptions of homogeneity of variance, and 2-way ANOVAs followed by Tukey Post Hoc tests were run with month and site as factors. Average concentrations of dissolved organic carbon [DOC (μM)], and dissolved nitrogen [TN (μM)] were compared among sites. The DOC concentrations could not be transformed to meet assumptions of normality, so a Kruskal-Wallis test was used followed by 2-sample Wilcoxon test pairwise comparisons with a sequential Bonferroni correction. A 2-way ANOVA was used followed by Tukey Post Hoc tests to compare TN concentrations with location (Panama or Belize) and site (reef or mangrove) as factors. Picoplankton composition and abundance were quantified at each site in terms of cell density, carbon content, and nitrogen content. Carbon and nitrogen content of prey items were estimated using the following literature conversion factors: heterotrophic bacteria, 0.02 pg C cell- 1 (Ducklow et al 1993) and 0.0038 pg N cell-1 (Caron et al. 1995); prochlorophytes, 0.05 pg C cell-1 (Morel et al. 1993) and 0.004 pg N cell-1 (Heldal et al. 2003); cyanobacteria, 0.47 pg C cell- 1 (Campbell et al. 1994) and 0.03 pg N cell-1 (Heldal et al. 2003); and picoeukaryotes, 0.8 pg C cell-1 (Verity et al. 1992) and 0.04 pg N cell-1 (Caron et al. 1995).

Data were transformed as necessary to meet assumptions of normality and homogeneity of variance. For each response variable, the total abundance of picoplankton was compared among sites using a 1-way ANOVA with Tukey Post Hoc tests. A 2-way ANOVA was also conducted followed by Tukey Post Hoc tests to compare the abundance of each prey type (heterotrophs, cyanobacteria, prochlorophytes, picoeukaryotes) at each site in order to understand the picoplankton community composition. Because the prochlorophyte and the picoeukaryote components contributed relatively little to the total picoplankton abundance, these prey types were combined with the cyanobacteria prey type to create a trophic group representing all autotrophs when the abundance of the autotrophic and heterotrophic components were compared across sites using a 2-way ANOVA with site and trophic group (autotrophs or heterotrophs) as factors, followed by Tukey Post Hoc tests.

Results

Summer light intensity measurements in Panama and Belize Mean daily light intensity measurements for Panama in August and for Belize in May ranged from 1,000 lux to 10,000 lux, with mangrove sites having the lowest light intensities and the highest light intensity occurring at the sea grass site (Fig 1.5a). The effect of site was significant for mean light intensity (Table 1.2). Mean light intensity at the Belize sea grass site was significantly higher than the Belize mangrove site (t = 7.114, p < 0.001) and all three Panama sites (R1: t = 3.991, p = 0.004; R2: t = 4.501, p < 0.001; M: t = 6.454, p < 0.001). Mean light intensity was not significantly different at the three reef sites (PAN-R1, PAN-R2, BZE-R: p > 0.321). While the mangrove sites in Belize and Panama had the lowest mean light intensities, only the mangrove site in Belize was significantly lower than the other Belize sites (BZE-R: t = 5.012, p < 0.001; BZE-SG: t = 7.114, p < 0.001). The mean light intensity at the mangrove site in Panama was not significantly lower than the two Panama reef sites (R1: t = 3.809, p = 0.079; R2: t = 2.559, p = 0.133).

Table 1.2 Panama and Belize Summer Light Intensity ANOVA Table Df Sum Sq Mean Sq F value p value Site 2 268198291 53639658 13.92 <0.0001 Residuals 35 134871523 3853472

Winter and summer light intensity measurements in Belize There was a significant effect of both month and site on mean light intensity (log) in Belize (Figure 1.5b; Table 1.3). Mean light intensities were significantly lower in December than in May (p = 0.002) and they were significantly lower at the mangrove than at the reef and sea grass sites (p < 0.0001).

Table 1.3 Belize December and May Light Intensity ANOVA Table Sum Sq Df F value p value Site 73.312 2 121.66 < 0.0001 Month 3.330 1 11.05 0.0016 Site x Month 0.379 2 0.63 0.5367 Residuals 16.270 54

Figure 1.5. (a) Mean summer light intensity in Panama and Belize and (b) mean light intensity in winter and summer in Belize. Letters indicate significance at α = 0.05. Sites are abbreviated as M = mangrove, R = reef, SG = sea grass. Error bars represent ± 1 s.e. of the mean.

Dissolved nutrient concentrations in Panama and Belize Mean DOC concentrations ranged from approximately 65 μM at the reef in Belize to 115 μM in the mangrove in Belize. DOC concentrations appear to vary more in the Belize mangrove than at the other sites. There was a significant effect of site on DOC concentration (Fig 1.6a; χ2 = 27.5744, df = 3, p < 0.0001). DOC concentrations were significantly higher in the Belize

mangrove (BZE-M) than at all other sites (PAN-M: p = 0.0002; PAN-R1: p = 0.0002; BZE-R: p = 0.003) and DOC concentrations in the Panama mangrove were the second highest (PAN-R1: p = 0.017; BZE-R: p = 0.0007). Finally, DOC concentrations on the Panama reef (PAN-R1) were significantly higher than on the Belize reef (BZE-R: p = 0.0006). Mean total dissolved nitrogen concentrations ranged from 1.4 μM on the Belize reef to 4.2 μM in the Belize mangrove. There was a significant interaction between location (Belize and Panama) and site (reef and mangrove) (Table 1.4; Fig 1.6b). The dissolved nitrogen concentrations were significantly higher in the Belize mangrove than all other sites (BDT-M: p = 0.0005; BDT-R1: p = 0.0001; BZE-R: 0.00002).

Table 1.4 Dissolved Nitrogen ANOVA Table Sum Sq Df F value p value Site 10.6361 1 12.1326 0.0013 Location 3.8612 4 4.4045 0.0429 Site x Location 15.5680 1 17.7586 0.0002 Residuals 31.5594 36

Figure 1.6. (a) Mean DOC and (b) mean TN concentrations at the mangrove and reef sites in Panama and Belize. Letters indicate significance at α = 0.05. Sites are abbreviated as M = mangrove, R = reef. Error bars represent ± 1 s.e. of the mean.

Picoplankton cell density, carbon, and nitrogen Mean density of picoplankton cells available to filter feeders at the different sites ranged from the highest densities occurring at the mangrove sites in Belize and Panama (~1,200,000 to

~1,500,000 cells/mL) to the lowest densities occurring at the reef sites in Belize and Curacao (~540,000 to ~640,000 cells/mL). There was a significant effect of site on picoplankton cell density (Table 1.5; Fig 1.7). When picoplankton cell density was partitioned by the four prey types, each differing in the amount of nutrition it represents to a filter feeder, there was also a significant interaction between site and prey type density (Table 1.6; Fig 1.7). At most sites, densities were ordered, from highest to lowest, as heterotrophs, cyanobacteria, prochlorophytes, than picoeukaryotes (p < 0.0001). Because the smallest and the largest of the autotrophic prey types contributed least to overall picoplankton densities, resources were also analyzed in terms of the relative abundance of each trophic group, heterotrophic and autotrophic cells. When the density of heterotrophic cells was compared to the autotrophic cells, there was a significant interaction between site and trophic group density (Table 1.7; Fig 1.7) and heterotroph cell density was significantly greater than autotroph cell density at every site (p < 0.0001).

Table 1.5 Total Picoplankton Cell Density ANOVA Table Df Sum sq Mean Sq F value p value Site 9 19.1777 2.13086 66.453 < 0.0001 Residuals 175 5.6115 0.03207

Table 1.6 Picoplankton Prey Type Cell Density ANOVA Table Sum sq Df F value p value Prey type 3523.0 3 13916.917 < 0.0001 Site 28.5 9 37.534 < 0.0001 Prey type x Site 312.1 27 136.987 < 0.0001 Residuals 60.6 718

Table 1.7 Picoplankton Trophic Group Cell Density ANOVA Table Sum sq Df F value p value Trophic group 626.73 1 14301.427 < 0.0001 Site 34.87 9 88.404 < 0.0001 Trophic group x Site 52.31 9 132.638 < 0.0001 Residuals 15.60 356

Figure 1.7. Mean (± 1 s.e) picoplankton cell density at each site, partitioned by prey type. Letters indicate significance at α = 0.05. No symbol: heterotrophs > cyanobacteria > prochlorophytes > picoeukaryotes. Symbol (*) indicates: heterotrophs > cyanobacteria = prochlorophytes > picoeukaryotes. In Curacao (†): heterotrophs > prochlorophytes > cyanobacteria > picoeukaryotes. (M=mangrove, R=reef, SG=seagrass).

When these cell densities were converted to the carbon they represent to a filter feeder, mean picoplankton carbon ranged from the highest levels of ~60,000 pg/mL to ~68,000 pg/mL in Panama to the lowest levels of ~24,000 pg/mL at the Belize and Curacao reef sites. There was a significant effect of site on picoplankton carbon (Table 1.8; Fig 1.8). There was a significant interaction between site and the carbon represented by each prey type (Table 1.9; Fig 1.8). Despite the fact that the density of heterotrophic cells was greater than the densities of all other cell types at all sites, cyanobacteria cells represented more carbon at most sites than heterotrophic cells. When the picoplankton carbon was divided into trophic groups, there was a significant interaction between site and trophic group (Table 1.10; Fig 1.8).

Table 1.8 Total Picoplankton Carbon ANOVA Table Df Sum sq Mean Sq F value p value Site 9 30.5267 3.3919 70.29 < 0.0001 Residuals 175 8.4447 0.0483

Table 1.9 Picoplankton Prey Type Carbon ANOVA Table Sum sq Df F value p value Prey type 1560.43 3 6164.367 < 0.0001 Site 28.50 9 37.535 < 0.0001 Prey type x Site 312.11 27 136.998 < 0.0001 Residuals 60.58 718

Table 1.10 Picoplankton Trophic Group Carbon ANOVA Table Sum sq Df F value p value Trophic group 5.736 1 114.956 < 0.0001 Site 65.191 9 145.155 < 0.0001 Trophic group x Site 43.721 9 97.351 < 0.0001 Residuals 17.765 356

Figure 1.8. Mean (± 1 s.e) carbon content of picoplankton at each site, partitioned by prey type. Letters indicate significance at α = 0.05. No symbol indicates: cyanobacteria > heterotrophs > picoeukaryotes > prochlorophytes. Symbol (*) indicates: heterotrophs > cyanobacteria > picoeukaryotes > prochlorophytes. Symbol (‡) indicates: cyanobacteria = heterotrophs > picoeukaryotes = prochlorophytes. Symbol (◊) indicates: heterotrophs > cyanobacteria = picoeukaryotes > prochlorophytes. In Curacao (†): heterotrophs > cyanobacteria = prochlorophytes > picoeukaryotes.

When picoplankton cell densities were converted into the nitrogen they represent to a filter feeder, values ranged from the highest levels of ~7,000 pg/mL in Panama to the lowest levels of ~2,000 pg/mL at the Belize sea grass site in December. There was a significant effect of site on picoplankton nitrogen (Table 1.11; Fig 1.9). There was a significant interaction between site and the abundance of nitrogen represented by each prey type (Table 1.12; Fig 1.9). At all sites, the heterotrophs contributed most to the picoplankton nitrogen. When the picoplankton nitrogen was separated by heterotrophic and autotrophic prey groups, there was a significant interaction between site and prey group (Table 1.13; Fig 1.9).

Table 1.11 Total Picoplankton Nitrogen ANOVA Table Df Sum sq Mean Sq F value p value Site 9 23.2391 2.58212 70.513 < 0.0001 Residuals 175 6.4083 0.03662

Table 1.12 Picoplankton Prey Type Nitrogen ANOVA Table Sum sq Df F value p value Prey type 2207.51 3 8693.285 < 0.0001 Site 28.96 9 38.022 < 0.0001 Prey type x Site 301.88 27 132.093 < 0.0001 Residuals 59.00 697

Table 1.13 Picoplankton Trophic Group Nitrogen ANOVA Table Sum sq Df F value p value Trophic group 72.179 1 1423.19 < 0.0001 Site 67.047 9 146.89 < 0.0001 Trophic group x Site 45.801 9 100.34 < 0.0001 Residuals 18.055 356

Figure 1.9. Mean (± 1 s.e) nitrogen content of picoplankton at each site, partitioned by prey type. Letters indicate significance at α = 0.05. No symbol indicates: heterotrophs > cyanobacteria > picoeukaryotes > prochlorophytes. Symbol (*) indicates: heterotrophs > cyanobacteria > picoeukaryotes = prochlorophytes. Symbol (‡) indicates: heterotrophs = cyanobacteria > picoeukaryotes = prochlorophytes. In Curacao (†): heterotrophs > cyanobacteria = prochlorophytes > picoeukaryotes.

Picoplankton abundance at reef and mangrove sites in Panama In Panama, the total picoplankton cell density at the mangrove site was significantly greater than the reef site (R2) located ~1 km away (Fig 1.10a; p < 0.01), but picoplankton density at the closer reef site (R1) was not significantly different from the mangrove site (Fig 1.10a; p > 0.05). When the picoplankton was partitioned into trophic groups, the cell densities for the heterotrophic and autotrophic components of the picoplankton were similar to the pattern of total picoplankton density (Fig 1.10a, b), except that the density of autotrophs at the mangrove site was not significantly different from either reef site (Fig 1.10b; p > 0.4). The carbon and nitrogen represented by picoplankton cells at R1 was significantly higher than R2 (carbon: Fig 1.10c; p < 0.01; nitrogen: Fig 1.10e; p < 0.004), but the picoplankton carbon and nitrogen at the mangrove site was not significantly different from either reef site (Fig 1.10c; p > 0.5). For the different picoplankton trophic groups, the autotrophic component represented significantly more carbon than the heterotrophic component at the Panama sites (Fig 1.10d; p < 0.0001). The pattern of just the autotrophic carbon component across sites was not different from the pattern of total

picoplankton carbon across sites (Fig 1.10c, d), but the heterotrophic carbon at the mangrove site was significantly greater than the heterotrophic carbon at the second reef site (R2) (Fig 1.10d; p = 0.03). The patterns of heterotrophic and autotrophic nitrogen mirrored the patterns of heterotrophic and autotrophic cell densities (Fig 1.10b, f).

Figure 1.10. (a) Mean picoplankton cell density at each Panama site, partitioned by prey type. (b) Mean cell density of heterotrophs and autotrophs at each Panama site. (c) Mean picoplankton carbon at each Panama site, partitioned by prey type. (d) Mean heterotrophic and autotrophic carbon at each Panama site. (e) Mean picoplankton nitrogen at each Panama site, partitioned by prey type. (f) Mean heterotrophic and autotrophic nitrogen at each Panama site. For all figures, letters indicate significance at α = 0.05 and error bars represent ± 1 s.e. of the mean.

Picoplankton abundance at reef and mangrove sites in Belize In Belize, the picoplankton densities at the mangrove site were significantly greater than the densities at the reef site (Fig 1.11a; p < 0.01). Picoplankton density was also significantly higher on the reef in December than in May (Fig 1.11a; p < 0.01). When picoplankton density is partitioned by trophic group, the heterotroph component was significantly greater at the mangrove site than the reef site in both months (Fig 1.11 b; p < 0.01), but the autotroph component was significantly greater at the reef site than the mangrove site in both months (Fig 1.11b; p < 0.0001). In terms of carbon represented by the picoplankton cells, the autotrophic component represented significantly more carbon than the heterotrophic component at the Belize reef site (Fig 1.11d; p < 0.0001). The opposite was true for the mangrove site (Fig 1.11d; p < 0.001). There was no significant difference in the carbon represented by autotrophs and heterotrophs at the sea grass site (Fig 1.11d; p > 0.05). The pattern of the autotrophic carbon component across sites was similar to the pattern of total picoplankton carbon (Fig 1.11c, d), except that there was significantly more autotrophic carbon at the reef site than the mangrove site in both December and May (Fig 1.11d; p < 0.0001). The pattern of the heterotrophic carbon component across sites was also similar to the pattern of total picoplankton carbon (Figure 1.11c, d), with the most heterotrophic carbon available at the mangrove site. The abundance of total picoplankton carbon at the reef site in December was largely represented by the autotrophic component, because the heterotrophic component at that site was not significantly different from the heterotrophic component of the picoplankton at the reef site in May (Fig 1.11d; p > 0.3). The pattern of heterotrophic nitrogen component was similar to the pattern of total picoplankton nitrogen sites (Fig 1.11), except that the heterotrophic nitrogen component was equivalent in May between the mangrove site and the sea grass site (Fig 1.11; p > 0.05). The pattern of the autotrophic nitrogen component across sites was similar to the pattern of autotrophic cell density because autotroph nitrogen was significantly higher at the reef site than the mangrove site in both December and May (Fig 1.11f; p < 0.0001) and did not vary between months at the sea grass site (Fig 1.11f; p = 1).

Figure 1.11. (a) Mean picoplankton cell density at each Belize site, partitioned by prey type. (b) Mean heterotroph and autotroph cell density at each site. (c) Mean picoplankton carbon at each Belize site. (d) Mean heterotroph and autotroph carbon at each site. (e) Mean picoplankton nitrogen at each site. (f) Mean heterotroph and autotroph nitrogen at each site. Letters indicate significance at α = 0.05. Sites are abbreviated as: M = mangrove, R = reef, SG = sea grass; error bars represent ± 1 s.e.

Picoplankton abundance at reef sites in Panama compared to Belize and Curacao The reef sites in Panama had significantly higher picoplankton cell densities, carbon abundances, and nitrogen abundances than the reef site in Belize (Fig 1.12; p < 0.05) and the reef site in Curacao (Fig 1.12; p < 0.01). The patterns of the heterotrophic and autotrophic components were similar to that of the total picoplankton cell density across sites (Fig 1.12a, b), except that autotrophic component at the Curacao reef site was not significantly different from one Panama reef site (R1) (Fig 1.12b; p > 0.05). The autotrophs represented significantly more carbon than the heterotrophs at the Panama and Belize reef sites (Fig 1.12b; p < 0.0001), but there was no significant difference at the Curacao reef site (Fig 1.12; p > 0.05). The heterotrophs represented significantly more nitrogen than the autotrophs at all reef sites (Fig 1.12 f; p < 0.0001). The patterns of the autotroph and heterotroph carbon and nitrogen components across reef sites were the same as the pattern of total picoplankton carbon abundance. Picoplankton abundance at mangrove sites in Panama and Belize The cell densities of the heterotrophic component were not significantly different from the total picoplankton cell densities at the mangrove sites in Panama and in Belize (Fig 1.13a; p > 0.6), but the cell density of the autotrophic component at the Panama mangrove was significantly higher than the autotrophic component at the Belize mangrove (Fig 1.13b; p < 0.0001). The carbon and nitrogen represented by the total picoplankton cells at the Panama mangrove site was significantly greater than at the Belize mangrove site (Fig 1.13c; p < 0.01; Fig 1.13e; p < 0.001) and that pattern was driven by the autotrophic component (Fig 1.13d; p < 0.0001; Fig 1.12f; p < 0.0001).

Figure 1.12. (a) Mean picoplankton cell density at each reef site, partitioned by prey type. (b) Mean cell density of heterotrophs and autotrophs at each reef site. (c) Mean picoplankton carbon at each reef site. (d) Mean heterotrophic and autotrophic carbon at each reef site. (e) Mean picoplankton nitrogen at each reef site. (f) Mean heterotrophic and autotrophic nitrogen at each reef site. For all figures, letters indicate significance at α = 0.05 and error bars represent ± 1 s.e. of the mean.

Figure 1.13. (a) Mean picoplankton cell density at each mangrove site, partitioned by prey type. (b) Mean heterotroph and autotroph cell density at each mangrove site. (c) Mean picoplankton carbon at each mangrove site. (d) Mean heterotroph and autotroph carbon at each mangrove site. (e) Mean picoplankton nitrogen at each mangrove site. Letters indicate significance at α = 0.05 and error bars represent ± 1 s.e.

Discussion

Light intensity and dissolved nutrients In general, the two mangrove sites in this study tended to have lower light intensity but higher dissolved nutrient concentrations than the reef and seagrass sites in Belize and Panama. (Fig 1.5 & 1.6). These dissolved nutrient and light patterns may have implications for the composition of the picoplankton community quantified at these sites. For instance, the Belize mangrove, with the lowest light intensity and the highest dissolved organic carbon and nitrogen concentrations, also had a greater relative abundance of carbon from heterotrophic cells (Fig 1.9), which do not depend on photosynthesis. Solar radiation has been previously shown to inhibit heterotrophic bacterial production to a depth of about 5 m (Visser et al. 1999), which may be one of the reasons heterotrophs are so dense in the mangroves, where there is lower light intensity. In addition to the free living microorganisms in the planktonic community, light intensity and dissolved nutrient patterns may influence the symbiotic relationships formed by sessile organisms with bacteria. In certain sponge species, a significant portion of biomass can be composed of symbiotic bacteria, both autotrophic and heterotrophic (Reiswig 1974). Some of these sponge species have been shown to obtain nutrition from grazing on picoplankton, nutrient transfer from heterotrophic and autotrophic symbionts, and significant removal of DOC (Reiswig 1981, de Goeij et al. 2008, Freeman and Thacker 2011, van Duyl et al. 2011, Strimaitis and Wulff in prep). These bacteriosponges may display plasticity in their trophic strategy depending on which strategy is optimal under different levels of picoplankton abundance, light intensity, and dissolved nutrients.

Picoplankton composition of water As predicted, picoplankton densities were greater at the mangrove sites in both Belize and Panama than the reef sites (Fig 1.10, Fig 1.11), but there was an exception to the pattern. The picoplankton densities at the Panama reef site in the sheltered bay closest to the developing coastline (PAN-R1) were equivalent to all mangrove sites. Picoplankton densities at site PAN- R1 were significantly greater than the picoplankton densities at the second reef site in Panama (PAN-R2), indicating that reef site densities were not similar mangrove densities at all reef sites in Panama, and there may be an important reason for this pattern. The main difference between these sites was that PAN-R1 is at the mouth of a sheltered bay that receives nutrient input from rapid coastal development and has less water flow than the other reef site. As predicted, both 25

Panama reef sites did have significantly higher picoplankton densities, picoplankton carbon abundance, and picoplankton nitrogen abundance than the Belize and Curacao reef sites (Fig 1.12). Although the picoplankton densities were not significantly different between the Panama mangrove site and the Belize mangrove site in May, the carbon and nitrogen available was greater in Panama than in Belize for the mangrove sites (Fig 1.13). Overall, these patterns support our initial predictions based on sponge growth rates that the mangrove sites in this study would have greater picoplankton resources than the reef sites, and the Panama sites in this study would have greater picoplankton resources than the Belize and Curacao sites. It is also important to understand the extent to which the different prey types contribute to picoplankton abundance because filter feeding animals may be limited by different prey. Furthermore, an excess of one prey type, like cyanobacteria, may actually become harmful to filter feeders (Peterson 2006). The picoplankton cell densities at all sites were dominated by heterotrophs, which are the smallest of the picoplankton prey types (Fig 1.7). At many sites, the heterotrophs also contributed most to the nitrogen available from picoplankton (Fig 1.9). While the autotrophic cells were much fewer in number, they are larger and contain more carbon and nitrogen per cell, which allowed the cyanobacteria prey type to represent the majority of the picoplankton carbon at most sites (Fig 1.8). In general, heterotrophic resources were higher at mangrove sites compared to reef sites included in this study (Fig. 1.10b, Fig 1.11b) and at Panama reefs compared to Belize and Curacao reefs included in this study (Fig 1.12b), while autotrophic resources were actually greater at reef sites than the mangrove sites (Fig. 1.10, Fig 1.11). These results signify that selectivity of filter feeders must be considered when making predictions about the relationships between resource levels and filter feeder distribution and abundance. For example, there is some evidence that some sponge species retain smaller cells more efficiently than larger cells (Reiswig 1971, Ribes et al. 1999). Different sites can have similar autotrophic carbon available to filter feeders, but completely different levels of heterotrophic carbon, which will matter to the success of an organism that selectively filters heterotrophic picoplankton. The distribution and abundance of many filter feeding groups may be indirectly controlled by differences in picoplankton abundance across sites, because picoplankton abundances can also influence trophic interactions within the complex microbial food web that includes multiple links between picoplankton, nanoplankton, algae larger than 5 μm, flagellates,

and ciliates in pelagic waters (reviewed by Sherr and Sherr 1991). While distinct picoplankton abundance patterns across sites may trickle up the trophic links, indirectly affecting many marine metazoans in these Caribbean ecosystems, only the distribution and abundance of filter feeders that specialize on efficiently clearing picoplankton will be directly affected. Sponges are unique because they specialize in efficiently retaining picoplankton as small as 0.1 μm with nearly 100% efficiency (Reiswig 1971, 1975, 1990, Ribes et al. 1999) and often dominate sessile animal biomass in Caribbean coral reef, mangrove, and sea grass ecosystems (Sutherland 1980, Wulff 2006, Biggs et al. in prep). Yet, studies on sponge filter feeding ecology are relatively rare compared to studies on other filter feeders, like bivalves. Despite this paucity of information and the persisting notion that sponges must be non-selective filter feeders, there is some evidence for selective retention of different particles in certain sponge species (Reiswig 1971, Turon et al. 1997, Ribes et al. 1999, Maldonado et al. 2010, Strimaitis and Wulff in prep). In light of this evidence, it is feasible that the distribution and abundance of some sponge species may be driven by bottom up forces because the abundances of different picoplankton prey types vary across the different sites compared in this study.

Conclusion Clearly is it important to know about the energy available in the pelagic system to filter feeding animals in the benthic system, because differences in available energy may fuel patterns of distribution, abundance, and growth rates in a bottom up fashion for these benthic animals. Only a handful of other studies have quantified picoplankton abundance in water flowing over filter feeders in coastal Caribbean environments where energy transfer from the pelagic to the benthic system is an important trophic process (Reiswig 1971, Reiswig 1981, Gast et al. 1998, Lesser 2006, Trussell et al. 2006, de Goeij et al. 2008). No study that we are aware of has compared picoplankton abundance across multiple sites in the Caribbean. Although we did not select the study sites specifically to test this variable, the pattern of higher picoplankton abundance at sites that receive more coastal nutrient input cannot be overlooked. While high picoplankton abundance can describe environments conducive for benthic filter feeders, the picoplankton group also contains disease causing microbes that may decimate certain species, removing their filtering power from the system (Wulff 2007). Previous work (Peterson et al. 2006) has demonstrated that when invertebrate filtering power is reduced, previously controlled phytoplankton populations can explode to detrimental levels, especially when fueled by coastal

inputs of nitrogen and phosphorous. Additional studies are necessary to investigate the filtering power currently present at sites with different picoplankton abundances and the potential of harnessing that power to preserve and restore water quality in these important ecosystems.

CHAPTER TWO

FILTER FEEDING RATES AND SELECTIVITY OF ERECT BRANCHING SPONGES ON CARIBBEAN CORAL REEFS

Introduction

Pelagic to Benthic Energy Transfer Filter feeding is a trophic strategy unique to aquatic environments because the physical properties of water allow organisms and particulate matter to be suspended in the water column. Filter feeding (used here synonymously with suspension feeding) is particularly dominant in benthic communities composed of animal groups that have evolved structures or morphologies adapted to utilize resources in suspension (Gili & Coma 1998). Passive filter feeders differ from active filter feeders because they depend on the natural flow of water to bring food in contact with their feeding structures, while active filter feeders pump water through a filtering device to separate food particles from the water so they can be concentrated and consumed (Riisgard and Larsen 2000). At the low energy cost of active filtering, sessile filter feeders can capture food items in bulk rather than individually via structures adapted to feeding in moving fluid environments (Hughes 1980, Riisgard and Larsen 1995). There are several main types of capture mechanisms in active filter feeding marine invertebrates, ranging from cirri trapping in bivalves to mucus net filter feeding in ascidians and gastropods to collar-sieving in sponges (Riisgard and Larsen 2000). Each mechanism differs in efficiency depending on the porosity of the filter, which sets a minimum size for particles that are efficiently retained from the water and can range from 5-10μm for large copepods down to 0.2μm for the collar cells in sponges (Jorgensen 1966). Zoobenthic filter feeders are ecologically important because they clear phytoplankton and bacteria from the water column and transfer energy from pelagic to benthic systems (Gili and Coma 1998, Lesser 2006, Peterson et al. 2006). The importance of benthic grazing is illustrated by evidence that sessile filter feeders regulate phytoplankton biomass in shallow waters like San Francisco Bay, Chesapeake Bay, and Florida Bay (Cloern 1982, Officer et al. 1982, Cooper and Brush 1993, Peterson et al. 2006). This ecosystem service is especially important in areas that receive high nutrient input, which can fuel damaging phytoplankton blooms. When filter feeders dominate the benthic biomass (e.g. oyster reefs and hard bottom sponge communities) and are

efficient at removing plankton, these organisms can have a dramatic effect on the ambient water. They can regulate water column properties either directly by grazing on members of the planktonic community or indirectly by intensifying nutrient cycling (Thorp and Casper 2003). Filter feeders also increase water clarity by removing particles, allowing more light to penetrate the benthos to photosynthesizing corals and sea grasses (Newell and Ott 1999, Peterson et al. 2006). Finally, selective feeding may enable filter feeders to influence the planktonic community structure of the overlying water column (Ward and Shumway 2004, Yahel et al. 2006). In shallow marine ecosystems, like bays and coral reefs, planktonic communities are largely shaped by substratum-current interactions and the activity of benthic communities (Riedl 1971, Sorokin 1994). Plankton in the near-bottom waters benefit from an increased flux of nutrients and organic matter from the benthos, but risk increased predation by filter-feeders (Pile et al. 1997, Gast et al. 1998, Yahel et al. 1998, van Duyl and Gast 2001, Scheffers et al. 2005). The planktonic community is dominated in biomass and productivity by members of the picoplankton (cells < 2μm) and nanoplankton (cells 2-20 μm) size classes, which include prokaryotic (cyanobacteria, prochlorophytes, and heterotrophic bacteria) and eukaryotic organisms (autotrophic and heterotrophic flagellates) (Platt et al. 1983, Stockner and Antia 1986, Burkill et al. 1993). Benthic filter feeders exploit different parts of the planktonic community depending on the porosity of their filtering mechanism, and understanding the trophic interactions between benthic and pelagic communities is an active area of research (Sherr and Sherr 1991, Van Wambeke et al. 1996). Knowledge of filtration rates and grazing selectivity for many members of benthic communities in shallow marine ecosystems, especially sponges, is incomplete and is necessary for understanding the influence of these organisms in structuring planktonic communities and for energy transfer from pelagic to benthic biomass.

Functional Importance of Sponges Sponges (Phylum Porifera) are one of the dominant filter feeding invertebrate groups occupying benthic space on Caribbean coral reefs, and at some locations sponge volume exceeds that of corals (Wulff 1994). On a shallow reef in San Blas, Panama, Wulff (2006) documented 35,000 cm3 of sponge biomass representing 1200 sponge individuals in a 12 m2 census area. On Looe Key reef at 7m depth in the Florida Keys, 100,000 cm3 of sponge biomass representing 1200 sponge individuals has been documented in a 16m2 census area (Biggs et al. in prep). The

sponge species on Caribbean coral reefs represent a variety of growth forms such as erect branching, massive, encrusting, and cryptic, but the erect branching morphology often contributes most to the total sponge volume on shallow Caribbean reefs (Wulff 2001, Strimaitis pers. obs.). Because sponges are the only filter feeding group dominating animal benthic biomass and efficiently removing picoplankton, bioremediation using sponges is currently a productive area of research, with many groups testing the potential of different species to restore polluted areas. Stabili et al. (2006) determined that Spongia officinalis var. adriatica, the most common Mediterranean ‘bath sponge’ that is commonly farmed off the coast of Italy, significantly decreased the bacterial concentration in ambient sea water, feeding preferentially on large and medium-sized bacteria. Peterson et al. (2006) quantified clearance rates of five dominant sponge species from Florida Bay of cultures of a common species of cyanobacteria, diatom, and dinoflagellate in laboratory experiments and of phytoplankton during in situ experiments. Finally, the effectiveness of filter feeder to mediate enrichment caused by aquaculture practices has also been tested (Angel et al. 2002, Zhang et al. 2009). Different sponge species display a variety of nutritional strategies, including taking up dissolved organic matter (Wilkinson and Garrone 1980, Reiswig 1981, Yahel et al. 2003, de Goeij et al. 2008), forming symbiotic associations with microorganisms (Wilkinson 1983, Diaz & Ward 1997, Hoffman et al. 2005, Freeman and Thacker 2011), and carnivory (Vacelet et al. 1995). However, sponges are most commonly known for their efficient removal of particles from the water column through filter feeding (Jorgensen 1966, Reiswig 1971, 1974; Gili et al. 1984, Wilkinson et al. 1984, Pile 1997, Ribes et al. 1999, Pile et al. 2003, Yahel et al. 2003), and the structure of the sponge body supports that these animals are well adapted to their primary trophic strategy (Fig 2.1). An extensive canal system pervades the sponge body and connects a network of chambers lined by choanocytes, flagellated cells that draw a unidirectional supply of ambient water in through the ostia (inhalant pores) and circulate the water throughout the sponge by the coordinated beating of their flagella. These choanocytes also have collars of microvilli, which act as sieves to capture particulate organic matter before ejecting cleared water and waste from the sponge through the osculum (exhalant opening) (Jorgensen 1966). Within this complex labyrinth of canals and chambers, almost all of the particles inhaled by the sponge come in contact with the pinacocytes lining the canals or the choanocytes in the filtering chambers, both of which have substantial phagocytic properties (Schmidt 1970, Wilkinson et al. 1984).

Figure2.1. Images of the internal canal architecture. (a) Cross section of a preserved Aplysina cauliformis under a dissecting microscope (40x). (b) Histological cross section of Aplysina fulva (400x). Arrow indicates water canal, (m) indicates the mesohyl, and (*) indicates feeding chamber lined by choanocytes.

Sponges are unique because they specialize in efficiently retaining picoplankton as small as 0.1 μm with nearly 100% efficiency (Reiswig 1971, 1975, 1990, Ribes et al. 1999). The picoplankton size class (0.1 to 2 μm) is one of the two groups mentioned previously that dominate the planktonic community and is also the size class containing pathogenic microbes, like bacteria, and microorganisms of ecological concern, like phytoplankton bloom species. The picoplankton that sponges ingest is often divided into four main prey types: picoeukaryotes, Synechococcus-type cyanobacteria, Prochlorococcus-type prochlorophytes, and heterotrophic bacteria (Ribes et al. 1999, Lesser 2006). From the point of view of a filter feeder, these prey types represent different amounts of nutrition in the forms of carbon and nitrogen (Table 2.1).

Table 2.1 Size, carbon, and nitrogen content of different picoplankton prey types Prey type Size (μm) C (pg cell-1) Reference N (pg cell-1) Reference Heterotroph ~0.6 ~0.02 Ducklow et al. 1993 ~0.0038 Caron et al. 1995 Prochlorophytes ~0.7 ~0.05 Morel et al. 1993 ~0.004 Heldal et al. 2003 Cyanobacteria ~1.1 ~0.47 Campbell et al. 1994 ~0.030 Heldal et al. 2003 Picoeukaryote ~1.5 ~0.80 Verity et al. 1992 ~0.040 Caron et al. 1995

While it is well established that many benthic suspension feeders, like bivalves, are selective in their grazing (Cranford and Gordon 1992, Ward and Shumway 2004), there is a persisting notion that sponges must be nonselective filter feeders. This idea stems from the presumption that the microvilli of the choanocytes act as a true sieve, with no mechanism for particle selection. Despite the little that is known about sponge feeding, there is some evidence for selective retention of different particle types by some sponge species. Reiswig (1971), studying three coral reef sponges, and Ribes et al. (1999), studying a Mediterranean sponge, both independently found that smaller bacteria are retained more efficiently than larger eukaryotic cells. Turon et al. (1997) fed latex beads of different sizes to 2 Mediterranean sponges and found the medium (1 μm) sized beads were retained most efficiently. Other sponge species have been reported to retain some pathogenic microbes more efficiently than others (Maldonado et al. 2010) and differentiate between “food type” bacteria and “symbiont type” bacteria (Wilkinson et al. 1984). Research on sponge filtering rates and selectivity is still in its preliminary stages as only a handful of sponge species have been studied and many of those species do not contribute significantly to sponge biomass in the study system. Most of what is known about sponge filter feeding efficiency and resource limitation comes from studies using one-pass sampling techniques that calculate retention efficiency by comparing the food available in the ambient water to the food remaining in the cleared water exhaled from the large central osculum of species with tube morphology (Reiswig 1971, Lesser 2006, Yahel et al. 2006). The majority of sponges contributing to biomass and numbers of individuals on Caribbean coral reefs have other growth forms (Wulff 2001), like erect branching morphology, that are not easily sampled with one pass techniques. Sponges pump water in proportion to their volume and, in order to understand how the majority of sponge biomass influences water quality on coral reefs, it is necessary to know something about the species that represent that biomass. When one pass sampling techniques are not appropriate, chamber experiments can be used to compare prey growth rates in control chambers to prey growth rates in experimental chambers with a sponge that is ingesting prey (Peterson et al. 2006, Ribes et al. 1999, Riisgard et al. 1993). The six coral reef species included in this study (Aplysina cauliformis and Aplysina fulva [Order: Verongida], Iotrochota birotulata and Desmapsamma anchorata [Order:

Poecilosclerida], and Amphimedon compressa and Niphates erecta [Order: Haplosclerida]. share a common erect branching morphology, but they are grouped in three separate orders that are each characterized by differences in tissue and skeletal composition. Skeletal structure and canal architecture may also influence sponge feeding ecology if they limit how quickly a volume of water can be moved through a sponge. Previous studies have also found that high microbial abundance sponges, like those in the genus Aplysina, have slower pumping rates, denser mesohyl, longer and narrower canals, and smaller choanocyte chambers than sponges without dense bacterial symbionts (Vacelet and Donadey 1977, Boury-Esnault et al. 1990, Weisz et al 2007). While the mechanisms governing feeding selectivity are unknown, the characteristics of water movement through a sponge species may influence whether those mechanisms can operate successfully. We predict that sponges with faster clearance rates will be less selective filter feeders than sponges with slower clearance rates. The diversity of these skeletal and tissue structures has also evolved under selection for mutualism between at least three of these species representing the three different orders. Growth rates and survival of Aplysina fulva, Iotrochota birotulata, and Amphimedon compressa are increased when two or more of these species adhere to one another and can benefit from the skeletal and chemical composition differences of their mutualistic partner that may protect against species specific predators or storm damage that they could not withstand individually (Wulff 1997). Based on the advantages of growing in close proximity, adhered to sponges of different orders, we predict that sponge species in different orders will partition the picoplankton prey types available on coral reefs, reducing competition for resources. This study is the first to quantify filtering rates for multiple species of erect branching sponges that contribute disproportionately to the sponge biomass at multiple Caribbean coral reef sites. We aimed to address three questions: 1) Do these sponge species effectively graze all four picoplankton prey types? 2) Do these sponge species filter selectively? 3) Do these sponge species remove prey at similar rates? These basic aspects of sponge filter feeding ecology are currently unknown and with this knowledge we can further address broader, ecosystem level questions such as: What is the effect on the water column of these species at documented densities on Caribbean reefs? How is the distribution and abundance of these species across Caribbean reefs controlled by the composition of the water column?

Methods

Study Sites This study was conducted on coral reefs in Panama and Belize. The experiments in Belize were conducted at a reef at a depth of 3m on the Blue Ground Reef near the Smithsonian station on Carrie Bow Cay and about 10 km from mainland Belize (site BZE; Fig 2.2a). In Panama, the study was conducted at two sites located in a shallow lagoon system directly off the coast and receiving high nutrient runoff from the nearby mountains and the developing town of Bocas del Toro. One study site (PAN-R1) is at a depth of 3 m across from the Smithsonian Tropical Research Institute at the opening of the bay to the town of Bocas del Toro. A second study site (PAN-R2) was at a depth of 6 m, near STRI point, which is about 1 km from the town of Bocas del Toro (Fig 2.2b). We conducted feeding experiments at these sites because they have different picoplankton abundances (Chapter 1) and correspond to differences in sponge growth rates (Wulff in prep). The Panama site closest to the town of Bocas del Toro (PAN-R1) has higher picoplankton abundance, in terms of both number of cells and volume of cells, than the Panama site at STRI point (PAN-R2), and both Panama sites have higher picoplankton abundances than the Belize site (BZE-R) (Chapter 1). At all three sites, the cyanobacteria and heterotroph prey types are significantly more abundant than prochlorophytes and picoeukaryotes (Chapter 1). The average temperature (30 oC) and light intensity (6000 lux) are not significantly different at the three sites (Chapter 1).

Figure 2.2. Study sites in (a) Belize and (b) Panama. Figure adapted from (a) Figure adapted from Smithsonian Institution CCRE Annual Report 2011 and (b) Google Maps.

Study Species The six sponge species used in this study are abundant on Caribbean coral reefs in both number and volume: Aplysina cauliformis and Aplysina fulva (Order Verongida), Iotrochota birotulata and Desmapsamma anchorata (Order Poecilosclerida), and Amphimedon compressa and Niphates erecta (Order Haplosclerida). Desmapsamma anchorata was only tested in Belize because it is not commonly found in Panama and Amphimedon compressa was only tested in Panama because the population in Belize had been decimated due to disease by 2005 (Wulff, pers. comm.). These species all have erect branching morphologies, but are grouped in three different taxonomic orders within the class Demospongiae that coincide with differences in their skeletal and chemical compositions. The Verongida are distinct from the other two orders because their skeletons are composed solely of spongin and do not contain spicules. The sponges from the genus Aplysina are considered “bacteriosponges” (Reiswig 1974), or sponge species with a significant biomass of microorganisms (Weisz 2006, Vicente 1990). The extent of nutrient transfer from these microbial symbionts to the sponge is currently an active area of research (Weisz et al. 2007, Fiore et al. 2010, Freeman and Thacker 2011).

Feeding Experiments To quantify sponge filtering rates and grazing selectivity we used in situ feeding experiments conducted on snorkel or SCUBA. These experiments were conducted by enclosing a fragment of each study species in a water tight chamber and comparing the change in the concentration of prey cells in the course of 20 minutes in a chamber with a sponge to the change in the concentration of prey cells in the course of 20 minutes in a control chamber without a sponge to calculate a clearance rate (adapted from Ribes et al 1999). Water samples were also collected from three or four control chambers (without a sponge) at each site. For each sponge species tested at each site, 10-12cm long fragments were cut from four to six different, healthy individuals and cable tied to 6 cm of ¼” PVC pipe on wire stakes (Fig 2.3a). The sponge fragments were allowed to heal and attach to the pipe, which generally required 3 to 5 days.

a. b. Figure 2.3. (a) Sponge fragments healing on PVC stands and (b) experimental chambers designed to quantify sponge filter feeding rates and selectivity.

The experimental chambers were designed from 6-L, food-safe, Rubbermaid containers with snap on lids (Fig 2.3 b). All experimental materials were conditioned, prior to use with animals, through sand scrubbing and soaking in ambient sea water for a minimum of 24 hours. To conduct each feeding experiment, the chamber was removed from the base and rinsed in ambient water while the sponge fragment was secured to the chamber base by the PVC stand. This method ensured that the sponge was never disturbed by human handling. Once the chamber was sealed, one 25-mL water sample was removed from the chamber at time points of 0 minutes, 10 minutes, and 20 minutes. At regular intervals throughout the experiment the chambers were gently swirled. Full syringes were removed from the chamber and replaced with empty syringes that had been rinsed 3 times in the ambient water. Full syringes were immediately transferred to a researcher in a boat on the surface and each syringe was mixed by inversion before 1.7 mL was filtered through a 100-μm mesh into a 2-mL cryovial and fixed with formaldehyde to a final concentration of 0.5%. Samples were stored on ice in a UV-proof box until the conclusion of the experiments, but no more than 8 hours. Samples were flash frozen in liquid nitrogen at the field station, transported back to Florida State University on dry ice, and stored at -80οF. The time points were chosen because the aim is to understand the effect of the sponge on the ambient water column rather than what a sponge does after it has depleted resources to levels much lower than its ambient norm or what the sponge does when it has been previously starved. Given the

size of the chamber and the size of the sponge fragments, twenty minutes is the maximum time that the water column in the chamber with a filtering sponge represented the ambient water column. The dimensions of each sponge fragment were measured in situ with a small ruler and calculated using geometric approximation of solids to normalize clearance rates for sponge size (as in Wulff 2001). Because each species has a unique skeletal composition, clearance rates were also standardized by ash free dry weight. Five fragments of each species were dried at 100oC for 24 h to determine the dry weight. Ash free dry weight (AFDW) was determined by combustion at 500oC for 6 hours (as in Ribes et al. 1999). Prior to combustion, sponges were rinsed to remove any salts. The relationship between ash (mg) and sponge volume (cm3) for each species was used to estimate ash free dry weights for sponge fragments that were not sacrificed. Sponges used in experiments were returned to stable substrata on the reef so growth rates could be measured over the subsequent year, although those data are not reported in this study.

Water Sample Analysis Water samples were analyzed using flow cytometry (see Chapter 1, methods) to quantify the concentration of each prey type (heterotrophic bacteria, Synechococcus-type cyanobacteria, Prochlorococcus-type prochlorophytes, and autotrophic picoeukaryotes).

Statistical Analysis All statistical analyses were conducted using R statistical software (R Developmental Core Team 2011). Depletion rates of prey in each chamber were calculated by assuming exponential growth and clearance of prey as described in Ribes et al. (1998). They prey growth rate (k) is calculated as follows:

where C0 and C1 are the prey concentrations in the chamber at the initial time t0 and at the final time t1. All the calculations in this study use the time points of = 0 min and = 20 min. The clearance rate R (volume swept clear biomass-1 time-1) is computed as follows:

where V is the volume of the chamber, b is the sponge biomass (mg AFDW) and g is the grazing coefficient (min-1), computed as:

where kc is the prey growth rate in the control chamber, and ka is the apparent growth in the chamber with the sponge. Finally, the ingestion rate, I (prey ingested biomass-1 time-1, is: where C is the average prey concentration during the experiment. The effect of grazing on each prey type was tested by comparing growth rates of prey in control (kc) and experimental (ka) chambers using a 3-way ANOVA with prey type, site, and species as fixed factors followed by Independent samples T tests or 2 sample Wilcoxon rank sum tests with sequential Bonferroni corrections for the post hoc multiple comparisons between each control and experimental chamber. To test for grazing selectivity, the percent of carbon and nitrogen in the total sponge diet contributed by each prey type was compared between the observed sponge diet measured in the chamber and the expected sponge diet measured in the ambient water measured at time zero. For a non-selective filter feeder, we would expect the observed diet to be similar to the expected diet (Ribes et al. 1999). Carbon and nitrogen content of prey items were estimated using literature conversion factors (Table 2.1). A MANOVA followed by Tukey Post Hoc tests for each dependent variable (% carbon and % nitrogen) were used to compare the observed diet to the expected diet. Site, species, and prey type were included as fixed factors. Additional ANOVA analyses with diet type (observed or expected), prey type (cyanobacteria, heterotrophs, picoeukaryotes, prochlorophytes), and site (if applicable) were conducted for Amphimedon compressa and Desmapsamma anchorata because they were not present at all three sites. Analysis of covariance (ANCOVA) was used to test for a significant effect of fragment size (mg AFDW) on estimated clearance rates (R) within a species. A separate ANCOVA was conducted for each prey group that was significantly grazed, and sponge species was included in the model as a fixed factor. A model simplification procedure was used to exclude non- significant variables and determine the minimal adequate model based on Akaike’s information criterion (Crawley 2007). Clearance rates were square root transformed to meet the assumptions of the test.

Results

Effect of grazing For prey growth in the control and treatment chambers, the main effects of prey type and species were both significant, and there was a significant interaction between prey type and sponge species, but not between prey type and site or prey type and site and species (Table 2.2; Fig 2.4). For the cyanobacteria and heterotroph prey types, all species significantly reduced the growth rate of the cyanobacteria compared to the control (Table 2.3; Fig 2.4). For the picoeukaryote prey type, none of the species significantly reduced the prey growth rate compared to the control (p > 0.05). For the prochlorophyte prey type, all species significantly reduced the prey growth rate compared to the control except the genus Aplysina (Table 2.3; Fig 2.4).

Table 2.2 Significance of Grazing ANOVA Table Sum Sq Df F value p value Prey type 0.0031 3 9.71 < 0.0001 Site 0.0011 2 5.38 0.0052 Species 0.0237 6 37.53 < 0.0001 Prey Type x Site 0.0007 6 1.08 0.3762 Prey Type x Species 0.0066 18 3.50 < 0.0001 Site x Species 0.0060 9 6.33 < 0.0001 Prey Type x Site x Species 0.0021 27 0.73 0.8345 Residuals 0.0248 235

Table 2.3 Pairwise Comparisons of Prey Growth in Control Chamber to Each Treatment Chamber cyanobacteria heterotrophs Prochlorophytes Sponge species W p value W p value W p value A. cauliformis 111 0.0009 115 <0.0001 NA > 0.05 A. fulva 139 <0.0001 135 <0.0001 NA > 0.05 A. compressa 90 0.0003 90 <0.0001 79 0.004 N. erecta 130 <0.0001 120 <0.0001 110 0.004 D. anchorata 24 0.009 24 0.01 24 .014 I. birotulata 124 0.0003 124 <0.0001 122 0.0001

Figure 2.4. Mean growth for each prey type in the control chamber and chambers with a sponge species present. Boxplots represent the range of values, with the middle 50% of the data enclosed in the box and the median indicated by the bold line. Symbols (*) indicate a significant difference between the treatment chamber and the control chamber for that prey type.

Selectivity of grazing To test for selectivity, the percent of carbon and nitrogen in the total diet contributed from each prey type was compared between the observed sponge diet measured in the chamber and the expected sponge diet measured in the ambient water. Site, species, and prey type were all fixed factors. The 4-way interaction was not significant, but the 3-way interactions for diet, prey type, and species and for diet, prey type, and site were significant (Table 2.4). Both 3-way interactions were significant for the percentage of carbon in the diet (Table 2.5), but none of the post hoc comparisons between the observed and expected diet for the same prey type at the same site were significant (p > 0.1). Only the 3-way interaction between diet, prey type, and species was significant for the percentage of nitrogen in the diet (Table 2.6). Separate analyses were conducted for A. compressa and D. anchorata because they were not present at all sites. For Desmapsamma anchorata there was a significant interaction between diet and prey type for both percent of carbon in the diet (Table 2.7) and nitrogen in the diet (Table 2.8).For Amphimedon

compressa, there was a significant effect of prey type (Table 2.9 and Table 2.10). The effect of observed or expected diet was not significant and the effect of site was also not significant.

Table 2.4 MANOVA Table Comparing Observed Sponge Diet to Expected Diet Based on Ambient Water Df Pillai F num Df den Df p value Diet type 1 0.0051 0.79 2 311 0.4529 Prey type 3 1.915 2333.55 6 624 <0.0001 Species 3 0.0004 0.02 6 624 0.9999 Site 2 0.0148 1.16 4 624 0.3257 Diet type x Prey type 3 0.2457 14.56 6 624 <0.0001 Diet type x Species 3 0.0003 0.02 6 624 0.9999 Prey type x Species 9 0.2533 5.03 18 624 <0.0001 Diet type x Site 2 0.0150 1.18 4 624 0.3200 Prey type x Site 6 0.2294 6.74 12 624 <0.0001 Species x Site 6 0.0033 0.09 12 624 0.9999 Diet type x Prey type x Species 9 0.4027 8.74 18 624 <0.0001 Diet type x Prey type x Site 6 0.1707 4.85 12 624 <0.0001 Diet type x Species x Site 6 0.0031 0.08 12 624 0.9999 Prey type x Species x Site 18 0.2577 2.56 36 624 <0.0001 Diet type x Prey type x Species x Site 18 0.1450 1.36 36 36 0.0834 Residuals 312

Table 2.5 ANOVA Table for Carbon in the Sponge Diet Df Sum Sq. Mean Sq. F value p value Diet type 1 33 33 0.5527 0.4578 Prey type 3 220820 73607 1244.2376 <0.0001 Species 3 3 1 0.0174 0.9969 Site 2 52 26 0.4421 0.6431 Diet type x Prey type 3 2187 729 12.3245 <0.0001 Diet type x Species 3 3 1 0.0174 0.9969 Prey type x Species 9 2349 261 4.4115 <0.0001 Diet type x Site 2 52 26 0.4407 0.6440 Prey type x Site 6 2146 358 6.0473 <0.0001 Species x Site 6 5 1 0.0139 0.9999 Diet type x Prey type x Species 9 5218 580 9.8002 <0.0001 Diet type x Prey type x Site 6 1138 190 3.2054 <0.0001 Diet type x Species x Site 6 5 1 0.0136 0.9999 Prey type x Species x Site 18 3737 208 3.5093 <0.0001 Diet type x Prey type x Species x Site 18 1771 98 1.6630 0.0448 Residuals 312 18457 59

Table 2.6 ANOVA Table for Nitrogen in the Sponge Diet Df Sum Sq. Mean Sq. F value p value Diet type 1 8 8 0.2064 0.6499 Prey type 3 290933 96978 2394.1753 <0.0001 Species 3 1 0 0.0088 0.9989 Site 2 7 4 0.0907 0.9133 Diet type x Prey type 3 635 212 5.2244 0.0016 Diet type x Species 3 1 0 0.0100 0.9986 Prey type x Species 9 2361 262 6.4760 <0.0001 Diet type x Site 2 7 4 0.0876 0.9161 Prey type x Site 6 1210 202 4.9782 <0.0001 Species x Site 6 2 0 0.0093 0.9999 Diet type x Prey type x Species 9 5163 574 14.1632 <0.0001 Diet type x Prey type x Site 6 402 67 1.6556 0.1315 Diet type x Species x Site 6 5 0 0.0088 0.9999 Prey type x Species x Site 18 3038 169 4.1673 <0.0001 Diet type x Prey type x Species x Site 18 1298 72 1.7804 0.0267 Residuals 312 12638 41

Table 2.7 ANOVA Table for Carbon in the Desmapsamma anchorata Diet Sum Sq. Df F value p value Prey type 19961.0 3 971.6766 <0.0001 Diet type 0.0 1 0.0001 0.9911 Prey type x Diet type 682.2 3 33.2067 <0.0001 Residuals 273.9 40

Table 2.8 ANOVA Table for Nitrogen in the Desmapsamma anchorata Diet Sum Sq. Df F value p value Prey type 31750 3 2489.799 <0.0001 Diet type 0 1 0.000 0.9944 Prey type x Diet type 345 3 27.055 <0.0001 Residuals 170 40

Table 2.9 ANOVA Table for Carbon in the Amphimedon compressa Diet Sum Sq. Df F value p value Prey type 46321 3 381.0404 <0.0001 Diet type 0 1 0.0003 0.9868 Site 0 1 0.0001 0.9915 Prey type x Diet type 43 3 0.3510 0.7886 Prey type x Site 132 1 1.0849 0.3630 Diet type x Site 0 1 0.0002 0.9882 Prey type x Diet type x Site 2 3 0.0123 0.9981 Residuals 2269 56

Table 2.10 ANOVA Table for Nitrogen in the Amphimedon compressa Diet Sum Sq. Df F value p value Prey type 48502 3 479.2788 <0.0001 Diet type 0 1 0.0000 0.9984 Site 0 1 0.0001 0.9924 Prey type x Diet type 44 3 0.4348 0.7289 Prey type x Site 302 3 2.9839 0.0388 Diet type x Site 0 1 0.0000 0.9982 Prey type x Diet type x Site 14 3 0.1392 0.93611 Residuals 1889 56

The post hoc tests indicated that, given the composition of the four prey types in the ambient water column, Aplysina cauliformis and Aplysina fulva ingested significantly more carbon and nitrogen from heterotrophs than expected (Fig 2.5; C: p < 0.0001, N: p < 0.001). In the order Haplosclerida, there were no significant differences between the observed carbon and nitrogen ingested from each prey type and the expected for Amphimedon compressa and Niphates erecta (Fig 2.5; p = 1). In the order Poecilosclerida, Iotrochota birotulata and Desmapsamma anchorata ingested significantly more nitrogen from cyanobacteria than expected (Fig 2.5; p < 0.002) and significantly less nitrogen from heterotrophs than expected (Fig 2.5; p < 0.0004). Desmapsamma anchorata also ingested significantly more carbon from cyanobacteria than expected (Fig 2.5; p < 0.0001), significantly less carbon than expected from heterotrophs (Fig 2.5; C: p = 0.016).

Figure 2.5. Comparison between the observed carbon and nitrogen contributed by each prey type to the sponge diet and the expected contribution based on the composition of the ambient water. (*) indicate significant difference between observed and expected diet at α = 0.05. † indicates significance at α = 0.1. Abbreviations for prey type represent: C = cyanobacteria, H = heterotrophs, Pi = picoeukaryotes, Pr = prochlorophytes. Error bars indicate ±1 s.e. of the mean.

Clearance rates An ANCOVA was conducted for each prey group that was effectively grazed to test for a relationship between sponge clearance rate (mL water AFDW-1 min-1) and sponge size (AFDW), with sponge species as a fixed factor. There was a significant negative relationship between cyanobacterial clearance rate (square root transformed) and sponge size (AFDW), and a significant effect of sponge species (Table 2.11, Fig. 2.6a). The interaction was not significant, indicating that the slopes of the lines are not different, but the slopes are. Amphimedon compressa cleared cyanobacteria significantly faster than all other species (p < 0.0059), and the two Aplysina species cleared cyanobacteria significantly slower than all other species (p < 0.0107).

Table 2.11 ANCOVA Table for Cyanobacteria Clearance Rate Df Sum Sq. Mean Sq. F value p value AFDW 1 0.761 0.761 34.58 < 0.0001 sponge species 5 1.026 0.205 9.33 < 0.0001 AFDW x sponge species 5 0.188 0.038 1.71 0.15 Residuals 55 1.211 0.022

There was also a significant negative relationship between heterotroph clearance rate (square root transformed) and sponge size (AFDW), and a significant effect of sponge species (Table 2.12, Fig. 2.6b). The interaction was not significant. Amphimedon compressa cleared heterotrophs significantly faster than all other species (p < 0.0023), and Niphates erecta cleared heterotrophs significantly faster than all species but Amphimedon compressa (p < 0.032).

Table 2.12 ANCOVA Table for Heterotroph Clearance Rate Df Sum Sq. Mean Sq. F value p value AFDW 1 0.671 0.671 48.38 < 0.0001 sponge species 5 0.762 0.152 10.97 < 0.0001 AFDW x sponge species 5 0.103 0.021 1.49 0.21 Residuals 54 0.749 0.014

Finally, there was a significant negative relationship between prochlorophytes clearance rate (square root transformed) and sponge size (AFDW), but no significant effect of sponge species and the interaction was also not significant (Table 2.13; Fig 2.6c).

Table 2.13 ANCOVA Table for Prochlorophyte Clearance Rate Df Sum Sq. Mean Sq. F value p value AFDW 1 0.162 0.1617 6.98 0.013 sponge species 3 0.036 0.0120 0.52 0.672 AFDW x sponge species 3 0.052 0.0174 0.75 0.531 Residuals 32 0.742 0.0232

Figure 2.6. Relationship between (a) cyanobacteria clearance rate and sponge size, (b) heterotroph clearance rate and sponge size, and (c) prochlorophyte clearance rate and sponge size. Symbols indicate species groupings: …..■..... = Amphimedon compressa; _.._▲_.._= Niphates erecta; ___ ●__ = Iotrochota birotulata and Desmapsamma anchorata (Poecilosclerida); _ _♦_ _ = Aplysina cauliformis and Aplysina fulva; ◊ = all species that significantly removed prochlorophytes (Amphimedon compressa, Niphates erecta, Iotrochota birotulata, Desmapsamma anchorata). Letters indicate significantly different intercepts at α = 0.05.

Discussion

Patterns of significant grazing, prey selectivity, and clearance rates varied by sponge species and taxonomic order, but reef site had no significant effect. Three of the four picoplankton prey types were significantly grazed compared to controls (Fig 2.4). None of the species significantly grazed picoeukaryotes, and Aplysina cauliformis and Aplysina fulva also did not significantly graze prochlorophytes. The picoeukaryotes and the prochlorophytes contribute least to the picoplankton abundance at these sites (Chapter 1), which makes it more likely that a sponge will capture enough of this prey to significantly reduce their densities in the ambient water column. Furthermore, it is possible that the picoeukaryotes were so rare that it was not possible to detect significant grazing in this study. Four of the six sponge species demonstrated selective filter feeding, defined as the composition of prey in the observed diet differing significantly from the composition of prey in the ambient water at time zero (Fig 2.5). Aplysina cauliformis and Aplysina fulva ingested more carbon and nitrogen from heterotrophs than expected given the composition of the ambient water, and Desmapsamma anchorata and Iotrochota birotulata ingested more carbon and nitrogen from cyanobacteria than expected. Overall, there was a negative relationship between clearance rate and sponge size (Fig 2.6), which has been previously recorded for other sponge species (Reiswig 1974, Frost 1980, Riisgard et al. 1993, Ribes et al. 1999). It has been suggested that this negative relationship may

be due to fewer choanocytes per biomass in larger sponges (Riisgard et al. 1993). Finally, not all sponge species removed prey at the same rate (Fig 2.6). Amphimedon compressa cleared cyanobacteria faster than the other species, while Aplysina cauliformis and Aplysina fulva cleared cyanobacteria slower than the other species, and both Haplosclerida species cleared heterotrophs significantly faster than the other species. Among the six sponge species tested in this study, it appears there may be order specific differences in their filter feeding ecology. In the order Verongida, Aplysina cauliformis and Aplysina fulva did not significantly clear the picoeukaryotes or the prochlorophytes from the ambient water and they selectively ingested the heterotrophic bacteria. A sponge species is classified as a high microbial abundance (HMA) sponge if it has microbial concentrations ≥108 cells g-1 (Hentschel et al. 2003). The Aplysina species are considered high microbial abundance (HMA) sponges (Weisz 2006, Vicente 1990), and these symbionts are able to transfer some nutrients to the sponge host (Freeman and Thacker 2011). This mutualism may reduce the dependence of these sponges on filter feeding as a primary trophic strategy and allow them to specialize on the heterotrophs, which are the most abundant prey type in number and have the smallest cell size, making them faster and easier to digest via phagocytosis. Aplysina cauliformis and Aplysina fulva also appear to clear prey at slower rates than Amphimedon compressa and Niphates erecta, the species that did not demonstrate selective filter feeding (Fig 2.5 and 2.6). Presumably, selective filter feeding would require greater handling time of prey and slower clearance rates, as observed in this study. The mechanisms for selective filter feeding in sponges are not yet known, although a study of a deep sea Hexactinellid sponge suggests that the encounter rate between inhaled particles and choanocytes may be low enough to allow for individual processing and selection of particles to ingest (Yahel et al. 2006). Previous studies have also found that HMA sponges have slower pumping rates, denser mesohyl, longer and narrower canals, and smaller choanocyte chambers than sponges without dense bacterial symbionts (Vacelet and Donadey 1977, Boury-Esnault et al. 1990, Weisz et al 2007). These differences suggest that Aplysina cauliformis and Aplysina fulva may have aquiferous systems that increase contact time between inhaled water and the microbial symbionts that can absorb dissolved nutrients and the phagocytic choanocytes that selectively filter the ambient picoplankton community. The two Poecilosclerida in this study, Iotrochota birotulata and Desmapsamma anchorata, significantly grazed all prey types except the picoeukaryotes, and ingested more of

their carbon and nitrogen from cyanobacteria than expected given the composition of the ambient water quantified at time zero (Fig 2.4, Fig 2.5). Iotrochota birotulata is categorized as an LMA species (Hentschel et al., unpublished data in Weisz et. al 2008). Desmapsamma anchorata is not reported as a HMA or LMA sponge, but has been shown to be able to utilize DOC in both coral reef and mangrove ecosystems (Hunting et al. 2010, van Duyl et al. 2011). If both species do not harbor significant microbial symbionts, it is assumed that they must rely on filter feeding as their primary trophic strategy. Therefore, it is intuitive that these species would selectively ingest the prey type that was not only abundant in number but also contained more carbon and nitrogen per cell digested than the other most abundant resource, the heterotrophs (Table 2.1). The clearance rates of cyanobacteria and heterotrophs for Iotrochota birotulata and Desmapsamma anchorata fell roughly in the middle of the Verongids and the Haplosclerids in this study (Fig 2.6), providing further evidence that selectivity may involve handling time that decreases clearance rates. Finally, the two Hapslosclerida in this study, Amphimedon compressa and Niphates erecta, had the fastest clearance rates of the most abundant prey types and did not demonstrate any filter feeding selectivity (Fig 2.5, Fig 2.6). These species are relatively free of microbial symbionts (Weisz 2006, Hentschel et al. unpublished data in Weisz et al. 2008) and rely on filter feeding to meet their nutritional needs. This strategy of clearing prey quickly may be optimal for these species because their tissue density is lower than many other Caribbean coral reef species (Weisz et al. 2008). Therefore, their skeletal structure is very porous, making it conducive to faster clearance rates that may not allow for the option of particle selection. Furthermore, Weisz et al. (2008) reported that there were significant density differences for Niphates erecta individuals collected from protected in shore and exposed outer reef habitats, with the less dense individuals inhabiting the protected areas, which suggests that differences in tissue density may be even greater between Niphates erecta and the other study species at protected sites like Bocas del Toro, Panama.

Implications Sponge feeding ecology is a complex story that is only starting to be told for the few species that have been studied. In 1997, Turon et al. remarked that the number of species whose feeding behavior had been investigated to date was too low to understand how it may be related to biological or structural elements, and the situation has not changed. It is not unreasonable to

expect that every sponge species that is explored will add another layer of complexity to what we know of the dynamics of sponge feeding. Yet this knowledge is imperative as this complexity is necessarily intertwined with the health of Caribbean coral reef ecosystems. If resources are partitioned across different sponge species as our study suggests, then diverse sponge communities are essential to maintain water quality by substantially removing all types of picoplankton. Although the picoplankton abundances on the three reefs included in this study were significantly different among sites (Chapter 1), there was no effect of site on clearance rate and selectivity for the members of a sponge species. This indicates that these species may not adjust how they feed as resources increase, but this lack of relationship needs to be tested with controlled genotype experiments. If this is the case, then more sponge biomass will be required to balance water quality at sites that receive excess nutrient runoff from coastal influences fueling phytoplankton blooms. Furthermore, if a sponge species is wiped out by a species specific disease, the unique role that species plays in maintaining water quality will also be lost. Species specific loss of sponges on coral reefs has been documented in several long term sponge census studies. Half of the most abundant species in a 14 year census study conducted on a shallow reef in San Blas, Panama were lost to disease (Wulff 2006). Amphimedon compressa had disappeared from reefs around Carrie Bow Cay, Belize by 2005 (Wulff, pers. comm.). Specific sponge species were lost to a cold water event in January 2010 from Looe Key Existing Management Area in the Florida Keys (Biggs et al. in prep). Finally, 75% of the sponge biomass documented on a census reef at the Blue Ground Range near Carrie Bow Cay, Belize was lost after a dense phytoplankton bloom in 2011 (Wulff in prep). The long term census data from Belize also revealed another incidence of dramatic species specific loss between the 2007 and the 2008 annual census, and the biomass had not recovered by the 2011 event (Wulff in prep). It is possible that the unprecedented dense and persistent phytoplankton bloom in 2011 could have been controlled before it reached damaging densities if the sponge biomass had been as great as it was 3 years earlier. The losses reported here likely only represent a small fraction of the actual sponge loss occurring on unmonitored reefs throughout the Caribbean. It is important to point out that the sponge losses from these events were only documented because they occurred in areas where a permanent census study was monitoring sponge individuals through time (as described in Wulff 2001). Sponges, unlike corals, do not leave evidence that they once existed in a coral reef community once they are gone, they fall apart and quickly vanish when they die.

Currently, sponges are rarely included in reef monitoring programs. When sponges are included, it is in terms of number of individuals or percent cover, which does not often allow for ecologically meaningful interpretations of influences on sponge distribution and abundance (Wulff 2012). Despite the sobering reality that large quantities of sponge filtering power have disintegrated from Caribbean coral reefs, the positive implication of this work is that knowledge on sponge filtration rates and selectivity can be used to inform bioremediation efforts. Bioremediation using sponges is not a new idea, but species specific information on clearance rates and selectivity should be taken into account. If sponge filter feeding ecology is understood in situ for species that represent significant biomass on reefs, supplemental sponge assemblages may be tailored to specific water quality issues by using the species with the most appropriate feeding behavior. Otherwise, we risk making mistakes such as directing extensive resources to attempting to improve water quality over reefs that are suffering from recurrent cyanobacteria blooms by supplementing the biomass of sponge species that retain heterotrophic bacteria most efficiently. This is the first study to quantify in situ feeding behavior at multiple sites for several sponge species that represent significant biomass in Caribbean coral reef ecosystems. These experiments were also conducted in areas where there are permanent census plots to follow individuals through time so that it will be possible to estimate the effect on the water column of these species at actual documented densities on Caribbean coral reefs. It is imperative that further studies are pursued to understand how all the common sponge species in Caribbean coastal ecosystems directly affect the water column. If information on sponge feeding in an area is evaluated in conjunction with data from long term monitoring of sponge individuals, extensive sponge losses can be documented and interpreted in terms of what those losses signify for the future water quality in that area.

CONCLUSION The functional roles and ecosystem services of filter feeding sponges are vital to the health of Caribbean coral reef ecosystems. We documented differences in the abundances of the four major picoplankton prey types across multiple Caribbean coastal environments, and also order specific differences in sponge filter feeding ecology. These two pieces of knowledge are necessary to understand how sponge distribution and abundance may be controlled by bottom up forces and how we can use this knowledge to inform bioremediation efforts. If resources are partitioned across different groups of sponges, then diverse sponge community assemblages are essential to maintain water quality. One of the major implications of this research is that long term monitoring of individual sponges on coral reefs is important in order to record changes in patterns of sponge distribution and abundance, which could be early indications of water quality declines that are possible to offset through bioremediation. Using sponges for bioremediation is not a new idea, but, with more information of sponge filter feeding ecology, it will be possible for species specific information to be taken into account. If feeding ecology is understood in situ for sponge species that represent significant biomass on reefs, supplemental sponge assemblages may be tailored to specific water quality issues by using the species with the most appropriate feeding ecology. These experiments were also conducted in areas where there are permanent census plots to follow individuals through time so that it will be possible to estimate the effect on the water column of these species at actual documented densities on Caribbean coral reefs. It is imperative that further studies are pursued to understand how all the common sponge species in Caribbean coastal ecosystems directly affect the water column. If information on sponge feeding in an area is evaluated in conjunction with data from long term monitoring of sponge individuals, extensive sponge losses can be documented and interpreted in terms of what those losses signify for the future water quality in that area.

REFERENCES

Angel, D. L., N. Eden, S. Breitstein, A. Yurman, T. Katz, and E. Spanier. 2002. In situ biofiltration: a means to limit the dispersal of effluents from marine finfish cage aquaculture. Hydrobiologia 469: 1-10.

Azam, F., T. Fenchel, J. G. Field, J. S. Gray, L. A. Meyerreil, and F. Thingstad. 1983. THE ECOLOGICAL ROLE OF WATER-COLUMN MICROBES IN THE SEA. Marine Ecology-Progress Series 10: 257-263.

“Bocas del Toro, Panama.” Map. Google Maps. Google, 1 April 2012. Web. 1 April 2012.

Boury-Esnault, N., L. De Vos, C. Donadey, J. Vacelet. 1990. Ultrastructure of choanosome and sponge classification. In: Rutzler K (ed) New Perspectives in sponge biology. Smithsonian Institution Press, Washington, D.C. pp 237-244.

Burkill, P. H., E. S. Edwards, A. W. G. John, and M. A. Sleigh. 1993. Microzooplankton and their herbivorous activity in the Northeastern Atlantic Ocean. Deep-Sea Research Part Ii- Topical Studies in Oceanography 40:479-493.Morris, Romma Heillig.

Campbell, L., Nolla, H.A., Vaulot, D. 1994. The importance of Prochlorococcus to community structure in the central North Pacific Ocean. Limnology and Oceanography 39: 954-960.

“Caribbean Sea.” Map. Google Maps. Google, 1 April 2012. Web. 1 April 2012.

Caron, D. A. and others 1995. THE CONTRIBUTION OF MICROORGANISMS TO PARTICULATE CARBON AND NITROGEN IN SURFACE WATERS OF THE SARGASSO SEA NEAR BERMUDA. Deep-Sea Research Part I-Oceanographic Research Papers 42: 943-972.

Cloern, J. E. 1982. DOES THE BENTHOS CONTROL PHYTOPLANKTON BIOMASS IN SOUTH-SAN-FRANCISCO BAY. Marine Ecology-Progress Series 9: 191-202.

Crain, J. “Manual for running samples on the Becton-Dickinson 4-Color Flow Cytometer in COAS.” Web manual 3 November 2003. Web. 6 January 2010.

Cole, J. J., S. Findlay, and M. L. Pace. 1988. BACTERIAL PRODUCTION IN FRESH AND SALTWATER ECOSYSTEMS - A CROSS-SYSTEM OVERVIEW. Marine Ecology- Progress Series 43: 1-10.

Cooper, S. R., and G. S. Brush. 1993. A 2,500-YEAR HISTORY OF ANOXIA AND EUTROPHICATION IN CHESAPEAKE BAY. Estuaries 16: 617-626.

Cranford, P. J. and D. C. Gordon. 1992. THE INFLUENCE OF DILUTE CLAY SUSPENSIONS ON SEA SCALLOP (PLACOPECTEN-MAGELLANICUS)

FEEDING-ACTIVITY AND TISSUE-GROWTH. Netherlands Journal of Sea Research 30:107-120.

Crawley, Michael J. The R Book. West Sussex: John Wiley & Sons, Ltd, 2007. Print. de Goeij, J. M., H. van den Berg, M. M. van Oostveen, E. H. G. Epping, and F. C. Van Duyl. 2008. Major bulk dissolved organic carbon (DOC) removal by encrusting coral reef cavity sponges. Marine Ecology-Progress Series 357:139-151.

Diaz, M. C. and B. B. Ward. 1997. Sponge-mediated nitrification in tropical benthic communities. Marine Ecology-Progress Series 156:97-107.

Ducklow, H. W., D. L. Kirchman, H. L. Quinby, C. A. Carlson, and H. G. Dam. 1993. Stocks and dynamics of bacterioplankton carbon during the spring bloom in the eastern North Atlantic Ocean. Deep-Sea Research 40: 245-263.

Fiore, C. L., J. K. Jarett, N. D. Olson, and M. P. Lesser. 2010. Nitrogen fixation and nitrogen transformations in marine symbioses. Trends in Microbiology 18:455-463.

Freeman, C. J. and R. W. Thacker. 2011. Complex interactions between marine sponges and their symbiotic microbial communities. Limnology and Oceanography 56:1577-1586.

Frost, T. M. 1980. CLEARANCE RATE DETERMINATIONS FOR THE FRESH-WATER SPONGE SPONGILLA-LACUSTRIS - EFFECTS OF TEMPERATURE, PARTICLE TYPE AND CONCENTRATION, AND SPONGE SIZE. Archiv Fur Hydrobiologie 90:330-356.

Gast, G. J., S. Wiegman, E. Wieringa, F. C. van Duyl, and R. P. M. Bak. 1998. Bacteria in coral reef water types: removal of cells, stimulation of growth and mineralization. Marine Ecology-Progress Series 167:37-45.

Gili, J. M., M. A. Bibiloni, and A. Montserrat. 1984. WATER-PUMPING ACTIVITY AND BACTERIAL RETENTION RATES IN-SITU OF 3 SPECIES OF LITTORAL SPONGES PRELIMINARY STUDY. Miscellania Zoologica (Barcelona) 8:13-22.

Gili, J. M. and R. Coma. 1998. Benthic suspension feeders: their paramount role in littoral marine food webs. Trends in Ecology & Evolution 13:316-321.

Heldal, M., D. J. Scanlan, S. Norland, F. Thingstad, and N. H. Mann. 2003. Elemental composition of single cells of various strains of marine Prochlorococcus and Synechococcus using X-ray microanalysis. Limnology and Oceanography 48: 1732-1743.

Hentschel, U., Fiesler, L., Wehrl, M., Gernert, C., Steinert, M., Hacker, J., Horn, M. 2003. Microbial diversity of marine sponges. In: Mueller WED (ed) Sponge (Porifera). Springer, Berlin Heidelberg New York.

Hoffmann, F., O. Larsen, V. Thiel, H. T. Rapp, T. Pape, W. Michaelis, and J. Reitner. 2005. An anaerobic world in sponges. Geomicrobiology Journal 22:1-10.

Hughes, R.N. 1980. Optimal foraging theory in the marine context, Oceanography and Marine Biology 18:423-481.

Hunting, E. R., J. M. De Goeij, M. Asselman, R. W. M. Van Soest, and H. G. Van Der Geest. 2010. DEGRADATION OF MANGROVE-DERIVED ORGANIC MATTER IN MANGROVE ASSOCIATED SPONGES. Bulletin of Marine Science 86: 871-877.

Jorgensen, C.B. 1966. Biology of Suspension Feeding. Pergamon Press, Oxford.

Koseff, J.R., J.K. Holen, S.G. Monismith, and J.E. Cloern. 1993. Coupled effects of vertical mixing and benthic grazing on phytoplankton populations in shallow, turbid estuaries. Journal of Marine Research 51: 843-868.

Lesser, M. P. 2006. Benthic-pelagic coupling on coral reefs: Feeding and growth of Caribbean sponges. Journal of Experimental Marine Biology and Ecology 328:277-288.

Maldonado, M., X. C. Zhang, X. P. Cao, L. Y. Xue, H. Cao, and W. Zhang. 2010. Selective feeding by sponges on pathogenic microbes: a reassessment of potential for abatement of microbial pollution. Marine Ecology-Progress Series 403:75-89.

Morel, A., Y. H. Ahn, F. Partensky, D. Vaulot, and H. Claustre. 1993. Prochlorococcus and Synechococcus: a comparative study of their optical properties in relation to their size and pigmentation. Journal of Marine Research 51: 617-649.

Newell, R.I.E., and J.A. Ott. 1999. Macrobenthic Communities and Eutrophication In: Malone, TC, NA Smadlaka, AMalej and LW Harding Jr. Ecology and ecosystem function of benthic communities in Chesepeake Bay and the Northern Adriatic Sea In. Land-Use Water quality and fisheries: a comparative ecosystem analysis of the northern Adriatic Sea and the Chesapeake Bay. Coastal and Estuarine comparisons series, American Geophysical Union.

Officer, C. B., T. J. Smayda, and R. Mann. 1982. BENTHIC FILTER FEEDING - A NATURAL EUTROPHICATION CONTROL. Marine Ecology-Progress Series 9:203-210.

Patten, N.L., A.S. Wyatt, R.J. Lowe, and A.M. Waite. 2011. Uptake of picophytoplankton, bacterioplankton and virioplankton by a fringing coral reef community (Nigaloo Reef, Australia). Coral Reefs 30: 555-567.

Peterson, B. J., C. M. Chester, F. J. Jochem, and J. W. Fourqurean. 2006. Potential role of sponge communities in controlling phytoplankton blooms in Florida Bay. Marine Ecology-Progress Series 328:93-103.

Pile, A. J., A. Grant, R. Hinde, and M. A. Borowitzka. 2003. Heterotrophy on ultraplankton communities is an important source of nitrogen for a sponge-rhodophyte symbiosis. Journal of Experimental Biology 206:4533-4538.

Pile, A. J., M. R. Patterson, M. Savarese, V. I. Chernykh, and V. A. Fialkov. 1997. Trophic effects of sponge feeding within Lake Baikal's littoral zone .2. Sponge abundance, diet, feeding efficiency, and carbon flux. Limnology and Oceanography 42:178-184.

Platt, T., D. V. S. Rao, and B. Irwin. 1983. PHOTOSYNTHESIS OF PICOPLANKTON IN THE OLIGOTROPHIC OCEAN. Nature 301:702-704.

R Development Core Team (2011). R: A language and environment for statistical computing. R Foundation for Statistical Computing,Vienna, Austria. ISBN 3-900051-07-0, URL http://www.R-project.org/.

Reiswig, H. M. 1971. PARTICLE FEEDING IN NATURAL POPULATIONS OF 3 MARINE DEMOSPONGES. Biological Bulletin 141: 568-591.

---. 1974. WATER TRANSPORT, RESPIRATION AND ENERGETICS OF 3 TROPICAL MARINE SPONGES. Journal of Experimental Marine Biology and Ecology 14: 231- 249.

---. 1975. BACTERIA AS FOOD FOR TEMPERATE-WATER MARINE SPONGES. Canadian Journal of Zoology-Revue Canadienne De Zoologie 53: 582-589.

---. 1981. Partial carbon and energy budgets of the bacteriosponge Verongia fistularis (Porifera: Demospongiae) in Barabados. Marine Ecology 2:273-293.

---. 1990. In situ feeding in two shallow-water Hexactinellid sponges. In: Rutzler K (ed) New Perspectives in sponge biology. Smithsonian Institution Press, Washington, DC, p 504- 510.

Ribes, M., R. Coma, M. J. Atkinson, and R. A. Kinzie. 2003. Particle removal by coral reef communities: picoplankton is a major source of nitrogen. Marine Ecology-Progress Series 257: 13-23.

Ribes, M., R. Coma, and J. M. Gili. 1998. Seasonal variation of in situ feeding rates by the temperate ascidian Halocynthia papillosa. Marine Ecology-Progress Series 175: 201-213.

---. 1999. Natural diet and grazing rate of the temperate sponge Dysidea avara (Demospongiae, Dendroceratida) throughout an annual cycle. Marine Ecology-Progress Series 176: 179- 190.

Riedl, R. 1971. WATER MOVEMENT ANIMALS. KINNE, OTTO (ED.). MARINE ECOLOGY. A COMPREHENSIVE, INTEGRATED TREATISE ON LIFE IN OCEANS AND COASTAL WATERS. VOL. I, PART 2. IX+1244P. ILLUS. MAPS.

WILEY-INTERSCIENCE, A DIVISION OF JOHN WILEY and SONS, LTD.: LONDON, ENGLAND; NEW YORK, N.Y., U.S.A:1123-1156.

Riisgard, H. U., and P. S. Larsen. 1995. Filter feeding marine macroinvertebrates – pump characteristics, modeling and energy cost. Biological Reviews of the Cambridge Philosophical Society 70: 67-106.

---. 2000. Comparative ecophysiology of active zoobenthic filter feeding, essence of current knowledge. Journal of Sea Research 44: 169-193.

Riisgard, H. U., S. Thomassen, H. Jakobsen, J. M. Weeks, and P. S. Larsen. 1993. SUSPENSION-FEEDING IN MARINE SPONGES HALICHONDRIA-PANICEA AND HALICLONA-URCEOLUS - EFFECTS OF TEMPERATURE ON FILTRATION-RATE AND ENERGY-COST OF PUMPING. Marine Ecology-Progress Series 96: 177-188.

Scheffers, S. R., R. P. M. Bak, and F. C. van Duyl. 2005. Why is bacterioplankton growth in coral reef framework cavities enhanced? Marine Ecology-Progress Series 299:89-99.

Schmidt, I. 1970. Phagocytose et pinocytose chez les Spongillidae. Zeitschrift fur Vergleichende physiologie 66: 398-420.

Sherr, E. B. and B. F. Sherr. 1991. PLANKTONIC MICROBES - TINY CELLS AT THE BASE OF THE OCEANS FOOD WEBS. Trends in Ecology & Evolution 6:50-54.

Smithsonian Institution. Caribbean Coral Reef Ecosystem Annual Report. Fort Pierce, FL: 2011.

Sorokin, Y.I. 1981. Microheterotrophic organisms in marine ecosystems. In: Longhurst, A. R. (ed.) Analysis of marine ecosystems. Academic Press, London, pp.293-342.

---. 1994. Coral Reef Ecology. Ecological Studies. Springer-Verlag, Berlin.

Stabili, L. and others 2006. Filtering activity of Spongia officinalis var. adriatica (Schmidt) (Porifera, Demospongiae) on bacterioplankton: Implications for bioremediation of polluted seawater. Water Research 40: 3083-3090.

Stockner, J.G. and Antia, N.J. 1986. Algal picoplankton from marine and freshwater ecosystems: a multidisciplinary perspective. Canadian Journal of Fisheries and Aquatic Science 43:2472-2503.

Sutherland, J. P. 1980. DYNAMICS OF THE EPIBENTHIC COMMUNITY ON ROOTS OF THE MANGROVE RHIZOPHORA-MANGLE, AT BAHIA DE BUCHE, VENEZUELA. Marine Biology 58: 75-84.

Thorp, J. H. and A. F. Casper. 2003. Importance of biotic interactions in large rivers: An experiment with planktivorous fish, dreissenid mussels and zooplankton in the St Lawrence River. River Research and Applications 19:265-279.

Trussell, G. C., M. P. Lesser, M. R. Patterson, and S. J. Genovese. 2006. Depth-specific differences in growth of the reef sponge Callyspongia vaginalis: role of bottom-up effects. Marine Ecology-Progress Series 323: 149-158.

Turon, X., J. Galera, and M. J. Uriz. 1997. Clearance rates and aquiferous systems in two sponges with contrasting life-history strategies. Journal of Experimental Zoology 278:22- 36.

Vacelet, J., N. Boury esnault, A. Fialamedioni, and C. R. Fisher. 1995. A METHANOTROPHIC CARNIVOROUS SPONGE. Nature 377:296-296.

Vacelet, J., and C. Donadey. 1977. ELECTRON-MICROSCOPE STUDY OF ASSOCIATION BETWEEN SOME SPONGES AND BACTERIA. Journal of Experimental Marine Biology and Ecology 30: 301-314.

Van Duyl, F. C. and G. J. Gast. 2001. Linkage of small-scale spatial variations in DOC, inorganic nutrients and bacterioplankton growth with different coral reef water types. Aquatic Microbial Ecology 24:17-26.

Van Duyl, F. C. and others 2011. Coral cavity sponges depend on reef-derived food resources: stable isotope and fatty acid constraints. Marine Biology 158: 1653-1666.

Van Wambeke, F., U. Christaki, and R. Gaudy. 1996. Carbon fluxes from the microbial food web to mesozooplankton. An approach in the surface layer of a pelagic area (NW Mediterranean Sea). Oceanologica Acta 19:57-66.

Verity, P. G., C. Y. Robertson, C. R. Tronzo, M. G. Andrews, J. R. Nelson, and M. E. Sieracki. 1992. RELATIONSHIPS BETWEEN CELL-VOLUME AND THE CARBON AND NITROGEN-CONTENT OF MARINE PHOTOSYNTHETIC NANOPLANKTON. Limnology and Oceanography 37: 1434-1446.

Vicente, V.P. 1990. Response of sponges with autotrophic endosymbionts during the coral- bleaching episode in Puerto Rico. Coral Reefs 8: 199-202.

Visser, P. M., E. Snelder, A. J. Kop, P. Boelen, A. G. J. Buma, and F. C. Van Duyl. 1999. Effects of UV radiation on DNA photodamage and production in bacterioplankton in the coastal Caribbean Sea. Aquatic Microbial Ecology 20: 49-58.

Ward, J. E. and S. E. Shumway. 2004. Separating the grain from the chaff: particle selection in suspension- and deposit-feeding bivalves. Journal of Experimental Marine Biology and Ecology 300:83-130.

Weisz, J.B. 2006. Measuring impacts of associated microbial communities on Caribbean reef sponges: searching for symbiosis. PhD thesis. University of North Carolina at Chapel Hill, N.C.

Weisz, J. B., U. Hentschel, N. Lindquist, and C. S. Martens. 2007. Linking abundance and diversity of sponge-associated microbial communities to metabolic differences in host sponges. Marine Biology 152:475-483.

Weisz, J. B., N. Lindquist, and C. S. Martens. 2008. Do associated microbial abundances impact marine demosponge pumping rates and tissue densities? Oecologia 155: 367-376.

Wilkinson, C.R. and R. Garrone. 1980. Nutrition in marine sponges. Involvement of symbiotic bacteria in the uptake of dissolved carbon. In: Smith, DC, Tiffon Y (eds) Nutrition in the lower Metazoa. Pergamon Press, Oxford, p 157-161.

Wilkinson, C. R. 1983. Net primary productivity in coral reef sponges. Science 219:410-412.

---. 1984. Immunological evidence for the Precambrian origin of bacterial symbioses in marine sponges. Proceedings of the Royal Society London Series B 220:509-517.

Wulff, J.L. 1994. Sponge-feeding by Caribbean angelfishes, trunkfishes, and filefishes. In: Soest RMW van, Kempen TMG van, Braekman J-C (eds) Sponges in time and space. Balkema, Rotterdam, pp265-271.

---. 1997. Mutualisms among species of coral reef sponges. Ecology 78: 146-159.

---. 2001. Assessing and monitoring coral reef sponges: Why and how? Bulletin of Marine Science 69:831-846.

---. 2006. Rapid diversity and abundance decline in a Caribbean coral reef sponge community. Biological Conservation 127: 167-176.

---. 2007. Disease prevalence and population density over time in three common Caribbean coral reef sponge species. Journal of the Marine Biological Association of the United Kingdom 87: 1715-1720.

---. 2012. Ecological interactions and the distribution, abundance, and diversity of sponges. In M. Becerro, M.Uriz, M. Maldonado, and X. Turon (Eds.), Advances in Marine Biology Volume 61: Advances in sponge science: Phylogeny, Systematics, Ecology (pp. 273 – 344). London: Elsevier.

Yahel, G., D. I. Eerkes-Medrano, and S. P. Leys. 2006. Size independent selective filtration of ultraplankton by hexactinellid glass sponges. Aquatic Microbial Ecology 45: 181-194.

Yahel, G., A. F. Post, K. Fabricius, D. Marie, D. Vaulot, and A. Genin. 1998. Phytoplankton distribution and grazing near coral reefs. Limnology and Oceanography 43: 551-563.

Yahel, G., J. H. Sharp, D. Marie, C. Hase, and A. Genin. 2003. In situ feeding and element removal in the symbiont-bearing sponge Theonella swinhoei: Bulk DOC is the major source for carbon. Limnology and Oceanography 48: 141-149.

Zhang, X., W., Zhang, L. Xue, B. Zhang, M. Jin, W. Fu. 2009. Bioremediation of bacteria pollution using the marine sponge Hymeniacidon perlevis in the intensive mariculture water system of Turbot Scophthalmus maximus. Biotechnology and Bioengineering 105: 59-68.

BIOGRAPHICAL SKETCH EDUCATION MSc student Biological Science, Florida State University, Tallahassee, FL (2008-present) Advisor: Dr. Janie Wulff Thesis Title: “Investigation of erect branching sponge filter feeding rates and selectivity on Caribbean coral reefs” Relevant Coursework: Population Ecology, Community Ecology, Evolution, Evolution of Ecological Symbiosis, Geostatistics, Quantitative Methods, Bioinformatics, Genome Sequencing & Analysis, Teaching Science to Maximize Learning and Retention, Assessment for Math and Science Educators

B.A. magna cum laude Middlebury College, Middlebury, VT (2003-2007) Thesis Topic: “Differential colonization of macrophytes with similar architecture by invertebrates and epiphytic algae in Vermont Lakes” Thesis Advisor: Dr. Sallie Sheldon

James Cook University, Townsville, QLD, Australia (Fall 2005) Relevant Coursework: Marine Invertebrate Biology (Dr. Gillian Brodie), Marine Conservation Biology (Dr. Geoff Jones)

PUBLICATIONS

Strimaitis, AM and SP Sheldon. 2011. A comparison of macroinvertebrate and epiphyte density and diversity on native and exotic complex macrophytes in three Vermont lakes. Northeastern Naturalist 18:149-160.

RESEARCH EXPERIENCE

Field Trips

2011 Bocas del Toro, Panama: conducted reef sponge feeding ecology experiments, investigated polychaete preference on mangrove sponge hosts, completed annual reef sponge census (August) The Florida Keys, FL: resurveyed 17m2 sponge census with Brendan Biggs (June) Gulf of Mexico Benthic Biodiversity Cruise: research cruise aboard the RV Weatherbird to document the “hidden biodiversity” of benthic habitats in the Gulf of Mexico (March)

2010 Bocas del Toro, Panama: conducted reef sponge feeding ecology experiments, investigated polychaete preference on mangrove sponge hosts, completed annual reef sponge census (August) CARMABI, Curacao: assisted Brendan Biggs with his reef restoration and sponge natural growth patterns research; conducted preliminary sponge feeding ecology experiment (July) The Florida Keys, FL: resurveyed 17m2 sponge census with Brendan Biggs (June) Carrie Bow Cay, Belize: carried out 2nd preliminary experiment investigating how sponges remove POC and DOC from the water; annual reef sponge census (May) The Florida Keys, FL: resurveyed 17m2 sponge census established in 2009 after the Cold Shock Event with Brendan Biggs (February)

2009 Carrie Bow Cay, Belize: carried out a pilot experiment sampling water for DOC and POC at time intervals from chambers enclosing filtering sponges (December) Bocas del Toro, Panama: Dr. Don Levitan and Dr. Nancy Knowlton’s research assistant for the annual Montastrea species complex coral spawn (September) Carrie Bow Cay, Belize: Dr. Janie Wulff’s research assistant for the annual sponge census in reef and sea grass habitats (July) The Florida Keys, FL: established a 17m2 sponge census with Brendan Biggs by identifying, enumerating, measuring, and mapping every sponge in the area (June)

Curatorial/Research Intern Reef HQ Aquarium, Townsville, QLD, Australia (2007-2008) Designed Saccoglossan sea slug breeding project Tested and maintained water quality in all aquaria (pH, DO, Redox, alkalinity, calcium, salinity, chlorine, ammonia) and bioassayed (Microtox, Artox) new intake water for toxins

Aquatic Ecology Research Assistant Sheldon Lab, Middlebury College (Summer ‘04 & ‘06) Raised biological control, Euhrychiopsis lecontei, for Eurasian Water Milfoil (Myriophyllum spicatum) and distributed over 10,000 weevils in Fairfield Lake, VT Sampled and manipulated algae for the Marine Biological Laboratory TIDE project Investigated competition between asexual and sexually reproducing macrophytes

Internet Digital Media Research Assistant Watters Lab, Middlebury College, VT (2006-2007) Searched scientific journals for supplemental movies on various cell biology topics Edited supplemental movies to create the professional figures in two published Cell Biology Education article series “Video Views and Reviews”

TEACHING EXPERIENCE

Assistant Coordinator for General Biology Lecture and Lab BSC1005 and 1005L Florida State University, Tallahassee, FL (Spring 2012 to present)

Saturday at the Sea (SATS) & Sea-to-See (C2C) Instructor Florida State University, Tallahassee, FL (Fall ’09 & ‘11, Spring ‘10)

General Biology Lecturer: Human Physiology Unit Florida State University, Biological Science, Tallahassee, FL (Summer ‘11)

Teaching Assistant Florida State University, Biological Science, Tallahassee,FL (2008-present) Experimental Marine Field Ecology (Fall ’11 Dr. D. Levitan) Animal Development Lab (Spring ’11 Dr. L. Keller) Experimental Biology Lab (Summer ‘09 & ‘10, Fall ‘09 Dr. L. Keller) Animal Diversity (Fall ‘08 & ‘10 Dr. W. Tschinkel, Spring ‘09 & ‘10 Dr. P. Spears) Animal Diversity Honcho (Fall ’10 C. Schultz)

Substitute Teacher in Sciences Medfield Public High School, Medfield, MA (2007) Taught AP Biology, Animal Physiology, Marine Biology, Chemistry, and Pre-Calculus Tutored students for math portion of the Massachusetts Comprehensive Assessment System

Teaching Assistant- Middlebury College, Biology Department, Middlebury, VT (2005-2007) Experimental Design and Analysis (under Dr. S. Sheldon) Cell Biology and Genetics (under Dr. V. Backus) Ecology (under Dr. R. M. Landis)

AWARDS, HONORS & RECOGNITIONS

Florida State University Outstanding Teach Assistant Award Nomination (2011) Horace Loftin Endowment (2010); $1000 for travel to Panama for Summer 2011 field work Departmental Citation for Excellence in Teaching (Spring 2010); Animal Diversity TA Horace Loftin Endowment (2009); $1000 for travel to Panama for Summer 2010 field work Lake Champlain Research Consortium Student Grant (2006); $500 Middlebury College Senior Work Fund (2006); $150

ORGANIZATIONS AND SOCIETIES

The Honor Society of Phi Kappa Phi Graduate Student Ecology and Evolution Reading and Discussion Group (EERDG) Financial Officer for academic year 2009-2010

POSTER PRESENTATIONS (PROFESSIONAL)

Evolution of form in Zoanthidea. Benthic Ecology Meeting, Mobile, AL (2011) Assessing change in sponge community composition after a disturbance: A case study of the Florida Keys 2010 Cold Shock. Benthic Ecology Meeting, Mobile, AL (2011) Cold shock event reinforces value of monitoring coral reef sponges. Linking Science to Management: A Conference and Workshop on the Florida Keys Marine Ecosystem, Duck Key, FL. (2010) Patterns of polychaete preference for mangrove sponge hosts. 8th International Sponge Conference, Girona, SPAIN. (2010) Mangrove sponge host preference of a surface dwelling polychaete. Benthic Ecology Meeting. University of North Carolina, Wilmington, NC. (2010)

ORAL PRESENTATIONS (PROFESSIONAL)

Filter feeding ecology of erect branching sponges on Caribbean coral reefs. Ecology and Evolution Seminar Series. Department of Biological Science, Florida State University, Tallahassee, FL. (2012)

Differential colonization of macrophytes with similar architecture by invertebrates and epiphytic algae in Vermont Lakes. Lake Champlain Research Consortium Student Symposium. Saint Michael’s College, Winooski, VT. (2007)

Differential colonization of macrophytes with similar architecture by invertebrates and epiphytic algae in Vermont Lakes. Biology and Molecular Biology and Biochemistry senior thesis presentations. Middlebury College, Middlebury, VT. (2007) SKILLS AND CERTIFICATIONS

Laboratory skills: flow cytometry, histology, total organic carbon analysis, PCR Computer Skills: R Statistical Software, SPSS, Linux OS, Wisconsin Package Genetics Computer Group, PAUP*, MS Excel, MS Word, Data Desk, QuicktimePro, Adobe Illustrator AAUS Scientific Diver Department of Biological Science Teach/Learning Workshop certification