THE EFFECTS OF HARD-BOTTOM HABITAT DEGRADATION

ON THE ECOLOGY AND BIOLOGY OF THE FLORIDA STONE CRAB

MENIPPE MERCENARIA IN THE FLORIDA KEYS

By

DEVON MICHAEL PHARO

A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF

FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF

MASTER OF SCIENCE

UNIVERSITY OF FLORIDA

2018

© 2018 Devon Michael Pharo

To my parents

4

ACKNOWLEDGMENTS

I would like to thank my parents, without whom I would not be the person I am today.

Growing up in the outdoors brought about a passion for nature and wildlife, which has lead me to

pursue marine science as a career. I would like to thank my wife for supporting me and helping

me with my studies over the past two years. I would also like to thank my advisor, Dr. Donald

Behringer, for supporting, guiding and helping me throughout my graduate studies at the

University of Florida. I also thank my committee members, Dr. William Lindberg and Dr.

William Patterson, for their guidance and knowledge on the subject matter as well as helping me

further my studies with new ideas. The endless encouragement and help from these individuals

throughout these past few years has made all the difference.

5

TABLE OF CONTENTS

page

ACKNOWLEDGMENTS ...............................................................................................................4

LIST OF FIGURES .........................................................................................................................7

LIST OF ABBREVIATIONS ..........................................................................................................9

ABSTRACT ...................................................................................................................................10

CHAPTER

1 BACKGROUND ....................................................................................................................12

2 THE EFFECTS OF HARD-BOTTOM HABITAT DEGRADATION ON

NUTRITIONAL CONDITION AND TROPHIC ECOLOGY OF MENIPPE

MERCENARIA ........................................................................................................................21

Introduction .............................................................................................................................21 Methods ..................................................................................................................................23

Objective 1: To Determine the Effects of Hard-Bottom Habitat Degradation on the

Nutritional condition of M. mercenaria. ......................................................................23 Objective 2: To Determine the Effects of Hard-Bottom Habitat Degradation on the

Trophic Ecology of M. mercenaria .............................................................................24 Results.....................................................................................................................................26

Objective 1: To Determine the Effects of Hard-Bottom Habitat Degradation on the

Nutritional Condition of M. mercenaria ......................................................................26

Objective 2: To Determine the Effects of Hard-Bottom Habitat Degradation on the

Trophic Ecology of M. mercenaria .............................................................................27 Discussion ...............................................................................................................................28

3 THE EFFECTS OF HARD-BOTTOM HABITAT DEGRADATION ON SITE AND

DEN FIDELITY OF MENIPPE MERCENARIA IN FLORIDA BAY ..................................46

Introduction .............................................................................................................................46 Methods ..................................................................................................................................49

Objective 1: To Determine the Effects of Hard-Bottom Habitat Degradation on the

Site and Den Fidelity of M. mercenaria. .....................................................................49

Objective 2: Response of M. mercenaria to Chemosensory Cues from the

Loggerhead Sponge, Spheciospongia vesparium. .......................................................50 Results.....................................................................................................................................52

Objective 1: To Determine the Effects of Hard-Bottom Habitat Degradation on the

Site and Den Fidelity of M. mercenaria. .....................................................................52

Objective 2: Response of M. mercenaria to Chemosensory Cues from the

Loggerhead Sponge, Spheciospongia vesparium. .......................................................52 Discussion ...............................................................................................................................53

6

4 CONCLUSIONS ....................................................................................................................66

LIST OF REFERENCES ...............................................................................................................69

BIOGRAPHICAL SKETCH .........................................................................................................79

7

LIST OF FIGURES

Figure page

1-1 Map of Florida Bay (USA) lying between the mainland of Florida and the Florida

Keys ...................................................................................................................................19

1-2 A large male Menippe mercenaria captured in Florida Bay in 2017.. ..............................20

2-1 Underside of a stone crab carapace with hepatopancreas being removed for analysis. ....32

2-2 Box plots of the weight to carapace width ratios of M. mercenaria between degraded

and non-degraded regions. .................................................................................................33

2-3 Box plots of the hemolymph refractive index of M. mercenaria between degraded

and non-degraded regions ..................................................................................................34

2-4 Box plots of the dry weight index of M. mercenaria between degraded and non-

degraded regions.. ..............................................................................................................35

2-5 Box plots of the dry weight index of M. mercenaria compared between males and

females. ..............................................................................................................................36

2-6 Box plots of the dry weight index of female and male M. mercenaria between

degraded and non-degraded regions. .................................................................................37

2-7 Weight versus carapace width for M. mercenaria samples (n=60) for which muscle

tissue was analyzed for 13C and 15N values at degraded and non-degraded sites. .........38

2-8 Dual isotope plot comparing δ13C and δ15N values of M. mercenaria between

degraded and non-degraded regions. .................................................................................39

2-9 Box plots of the δ13C and δ15N values of M. mercenaria between degraded and non-

degraded regions. ...............................................................................................................40

2-10 Dual isotope plot depicting δ13C and δ15N values between male and female M.

mercenaria. ........................................................................................................................41

2-11 Box plots of the δ13C and δ15N values of M. mercenaria between females and males .....42

2-12 Dual isotope plot depicting δ13C and δ15N values of M. mercenaria between size

classes. ...............................................................................................................................43

2-13 Box plots of the δ13C and δ15N values of M. mercenaria between size classes. ...............44

2-14 Dual isotope plot comparing δ13C and δ15N values for multiple species between

degraded and non-degraded habitat. ..................................................................................45

3-1 Common dens of M. mercenaria in hard-bottom habitat ..................................................57

8

3-2 Degraded and non-degraded sites of Florida Bay used in fidelity study ...........................58

3-3 Tags and weights used to mark individual crabs ...............................................................59

3-4 Sponges and Y-mazes used in chemosensory trials. ..........................................................60

3-5 Mean percentage of crabs recaptured within their original dens over the course of 8 d

in degraded and non-degraded sites. ..................................................................................61

3-6 Mean percentage of crabs recaptured on sites over the course of 8 d in degraded and

non-degraded sites. ............................................................................................................62

3-7 Fidelity of female crabs to individual dens and sites over the course of 8 d in

degraded and non-degraded sites. ......................................................................................63

3-8 Fidelity of male crabs to individual dens and sites over the course of 8 d in degraded

and non-degraded sites. ......................................................................................................64

3-9 Results of the final selections of M. mercenaria in chemosensory trials. .........................65

9

LIST OF ABBREVIATIONS

ANOVA Analysis of Variance

C Carbon

CW Carapace Width

DWI Dry Weight Index

GPS Global Positioning System

HABs Harmful Algal Blooms

N Nitrogen

P Phosphorus

PSP Paralytic Shellfish Poisoning

RI Refractive Index

10

Abstract of Thesis Presented to the Graduate School

of the University of Florida in Partial Fulfillment of the

Requirements for the Degree of Master of Science

THE EFFECTS OF HARD-BOTTOM HABITAT DEGRADATION

ON THE ECOLOGY AND BIOLOGY OF THE FLORIDA STONE CRAB

MENIPPE MERCENARIA IN THE FLORIDA KEYS

By

Devon Michael Pharo

August 2018

Chair: Donald C. Behringer

Major: Fisheries and Aquatic Sciences

The stone crab, Menippe mercenaria, supports one of the most economically important

fisheries in the southeastern United States. Hard-bottom habitat in the Florida Keys is an

important habitat for M. mercenaria and is characterized by large sponges, octocorals, and

macroalgae on a porous limestone bottom. Cyanobacteria blooms, consisting primarily of

Synechococcus sp., have periodically occurred in Florida Bay for at least the past several

decades, resulting in mass sponge mortalities, most notably the loss of loggerhead sponges.

Juvenile and young adult M. mercenaria are predominantly found residing in loggerhead

sponges or solution holes. I examined the potential impacts of sponge loss and overall habitat

degradation by comparing stone crab nutritional condition, trophic position, and site fidelity

between degraded and non-degraded regions of the Bay. I also investigated whether M.

mercenaria potentially use chemical cues from sponges to navigate in hard-bottom habitat.

Menippe mercenaria from degraded and non-degraded habitats were of similar nutritional

condition regardless of index used. Using stable isotope analysis, I found M. mercenaria from

degraded versus non-degraded habitats to be feeding at the same trophic level, but the carbon

sources supporting crabs in these regions differed significantly. This difference could be

11

attributed to a shift in prey choice or a shift in the isotopic signatures of primary producers

within the system. Menippe mercenaria could also be foraging more opportunistically within

degraded regions of Florida Bay. Stone crabs were attracted to chemical cues from loggerhead

sponges over seawater alone, indicating that they may indeed use cues exuded by sponges to

navigate hard-bottom and perhaps reduce the time to locate shelter.

The impact of habitat degradation caused by cyanobacterial blooms on this important

benthic species appears to be minimal. Menippe mercenaria maintained a similar nutritional

condition despite a shift in trophic niche, indicating acclimation to their altered habitat through a

shift in diet. Site and den fidelity remained similar regardless of the loss of loggerhead sponges;

probably due to the abundance of solution holes available as shelters throughout hard-bottom.

Overall, results indicate a high resilience to this particular natural disturbance by M. mercenaria.

12

CHAPTER 1

BACKGROUND

Anthropogenic impacts to ecosystems are increasing globally as the human population of

our planet continues to rise. As we continue to burn fossil fuels, CO2 levels have risen

exponentially leading to an increase in atmospheric and ocean temperatures. These changes,

among others, are expected to magnify the frequency and intensity of algal blooms into the

future (Aquino-Cruz et al. 2018, Paerl and Huisman 2008, Moore et al. 2008). Harmful algal

blooms, otherwise known as “HABs”, are of particular concern, not only affecting human health

(Heisler et al. 2008), but also damaging important ecosystems (Tiling and Proffitt 2017) and the

services they provide. The phytoplankton that make up HABs, such as cyanobacteria,

dinoflagellates, and diatoms (Glibert et al. 2005), are naturally occurring in freshwater and

marine environments, but manifest into blooms when stimulated and fueled by a nutrient supply,

which occurs naturally (Anderson et al. 2002) or by anthropogenic means (Fogg 1969, Paerl

2011, Reynolds 1987). Excess nutrient loading, or eutrophication, is the enrichment of a body of

water and results from agricultural, wastewater, and industrial runoff (Anderson et al. 2002,

Vitousek et al. 1997). The two major nutrients that typically fuel algal blooms, nitrogen and

phosphorus, are normally the limiting macronutrients within freshwater or marine systems

(Howarth 2008, Paerl et al. 2011). Harmful algal blooms can be grouped into two categories,

toxic and non-toxic. Toxic algal blooms can cause serious threats to humans and wildlife through

inhalation or ingestion of contaminated water or shellfish (Carmichael 2001, Ibelings et al.

2014). At high volumes, non-toxic blooms can also be detrimental to aquatic systems, depleting

oxygen levels and blocking sunlight to the benthos which can devastate benthic habitat, causing

further harm to aquatic organisms (Anderson et al. 2002).

13

The adverse effects and implications of HAB’s in aquatic environments have become

increasingly recognized as the frequency and intensity of blooms have increased. Many

economically and ecologically important species, as well as the communities they help support,

have been affected by these blooms. For example, in 1998, the dinoflagellate, Ostreopsis cf.

ovata, caused a major die off of the sea urchin, Echinometra lucunter, in Rio de Janeiro (Ferreira

2006). This dinoflagellate also was shown to affect fertilization and early development of

another sea urchin, Lytechinus variegatus, along the coast of Brazil (Neves et al. 2018). Another

toxic dinoflagellate, Alexandrium excavatum, was shown to induce heavy mortality in finfish

larvae, including the commercially important American mackerel in the Gulf of St. Lawrence,

Canada (Robineau et al. 1991). Harmful algal blooms can also affect benthic habitat directly,

causing indirect effects on species that depend on it. In the mid 1980’s, a brown tide of

Aureococcus anophagefferens occurred in several bays along the mid-Atlantic. This led to high

mortalities of eelgrass, Zostera marina, increasing light absorption within the water column,

which in turn caused recruitment failure of the commercially valuable bay scallop, Argopecten

irradians (Bricelj et al. 1987, Cosper et al. 1987, Dennison et al. 1989). In Narragansett Bay,

Rhode Island, the same bloom forming species led to reproductive failure and high mortalities in

the blue mussel, Mytilus edulis (Tracey 1988). Impacts such as these can be seen around the

globe affecting both habitat and organisms, which can further harm major fisheries and tourism.

For example, the high frequency of HABs experienced in Florida has continued to have major

implications on the environment and local economy (Larkin and Adams 2007).

With nearly 21 million people living in Florida, anthropogenic factors can play a large

role in the high frequency of HAB’s throughout the state. High agricultural, industrial, and

sewage runoff into Florida’s many water bodies facilitated by heavy rainfall contribute high

14

amounts of nutrients into systems leading to an increase in the frequency of algal blooms (Hu et

al. 2004, Lapointe and Bedford 2010, Phlips and Badylak 1999). The dinoflagellate, Karenia

brevis, also known as red tide, occurs on the Gulf coast of Florida almost yearly. This species is

associated with neurotoxic shellfish poisoning (NSP) which can be obtained through ingestion of

brevetoxins in contaminated shellfish (Backer 2009, Steidinger 1993). Karenia brevis also

produces aerosolized brevetoxins that can be inhaled by humans and wildlife causing respiratory

issues (Kirkpatrick et al. 2004). High mortalities have been reported in fish, manatees and

double-crested cormorants along the coasts of the Gulf of Mexico due to these highly toxic

blooms (Bossart et al. 1998, Kreuder et al. 1998, Trainer and Baden 1999). Karenia brevis has

also been shown to lower food consumption and decrease survivorship among Florida stone

crabs, Menippe mercenaria, on the west coast of Florida (Gravinese et al. 2018).

One group of HAB-causing phytoplankton, the cyanobacteria, are widely responsible for

effecting marine and estuarine communities (Paerl and Huisman 2005, Paerl et al. 2011). In

2006, the cyanobacterium, Lyngbya majuscule, bloomed in the Indian River Lagoon. Shading

from the bloom decreased the density of Halodule wrightii, a common seagrass, as well as the

density of a common bivalve, Macoma constricta (Tiling and Proffitt 2017). Cyanobacteria have

been found to produce the neurotoxin beta-N-methylamino-L-alanine (BMAA) which can

biomagnify in aquatic food webs and cause neurodegenerative diseases such as Parkinson’s and

Alzheimer’s (Brand et al. 2010). Harmful BMAA concentrations were found in species

consumed by humans, such as pink shrimp and blue crabs from Biscayne Bay, Florida. Another

cyanobacterium, Lyngbya sp., is a benthic form that often blooms on the coral reef tract in south

Florida and can negatively impact corals and other invertebrates (Paul et al. 2005). Non-toxic

15

cyanobacteria such as Synochococcus sp. can also have deleterious effects on habitat, such as has

occurred in Florida Bay in recent decades.

Florida Bay, which lies between the mainland of Florida and the Florida Keys (Fig. 1-1),

is well known for its productivity, biodiversity, and ecological importance as a nursery habitat

for many species of invertebrates and fish (Homquist et al. 1989, Thayer and Chester 1989,

Zieman et al. 1989). Seagrass beds, primarily made up of Thalassia testudinum, cover

approximately 60-70% of the Bay (Zieman et al. 1989) providing habitat and foraging grounds

for many species. Another 30-40% of the Bay consists of hard-bottom habitat (Behringer and

Butler 2006, Bertelson et al. 2009), an integral marine feature of the Florida Keys. Hard-bottom

habitat is characterized by sponges, solitary hard corals, octocorals, and patches of macroalgae

on a porous limestone bottom covered in a thin layer of sediment (Butler et al. 1995, Behringer

and Butler 2006). A diversity of high-density sponges is synonymous with this bottom, which

helps create its complex three dimensional structure. Loggerhead sponges, Speciospongia

vesparium, are the largest and most abundant of the sponge species (Peterson et al. 2006) and are

used as shelter by many benthic invertebrates, such as the Caribbean spiny lobster, Panulirus

argus (Butler et al. 1995) and Florida stone crab, Menippe mercenaria (Behringer and Hart

2017). Many other smaller benthic organisms, such as snapping shrimp, live within the internal

channels of this sponge as well (Pearse 1950).

Over the past several decades, Florida Bay has been affected by a series of cyanobacteria

blooms, dominated mainly by the picoplankton sized cyanobacterium, Synechococcus sp., which

has drastically changed the ecology of the Bay north of the middle Keys (Butler et al. 1995,

Phlips and Badylak 1996, Wall 2012). Over the past century, the natural flow of freshwater from

the Everglades into Florida Bay has been altered, leading to changes in the community structure

16

of both ecosystems. Nutrient loading from Taylor Slough combined with sediment resuspension

in the north central region of the Bay are responsible for elevated levels of cyanobacteria blooms

throughout the year (Phlips and Badylak 1996) and wind events often push these blooms

southwards towards the Florida Keys. The network of shallow mud banks throughout the north

central region of the Bay also contribute to the longevity of blooms with very low water

exchange with the open sea (Phlips et al. 1999). These blooms have coincided with massive

sponge mortalities in the northeast region of the Bay that have killed over 40% of loggerhead

sponges and over 70% of other sponge species (Butler et al. 1995, Butler et al. 2015). The loss of

loggerhead sponges in impacted regions has led to a dramatic decrease in available shelters and

overall habitat for many benthic species. In particular, these blooms have had negative effects on

the distribution and abundance of the Caribbean spiny lobster (Butler et al. 1995). Another

ecologically and economically important species that may be impacted by these blooms is the

Florida stone crab, M. mercenaria, but data are currently lacking to discern the effects of sponge

loss and habitat degradation on stone crabs in this system.

The Florida stone crab, M. mercenaria, and the Gulf stone crab, Menippe adina, occur

throughout the southeastern United States and Caribbean. Menippe mercenaria ranges from

Florida to North Carolina, also occurring throughout the Caribbean, Yucatan Peninsula and

Belize (Ong and Costlow 1970, Say 1818). Menippe adina occurs westward from northwestern

Florida along the Gulf of Mexico coast into Mexico (Williams and Felder 1986). The stone crab

is the largest xanthid crab within its range (Lindberg and Marshall 1984) and is characterized by

a pair of large chelipeds which account for over 50% of the overall weight of the crab (Fig. 1-2).

Stone crabs are almost entirely carnivorous and use their large crusher claw to break open and

feed on bivalves and gastropods (Menzel and Nichy 1958, Powell and Gunter 1968, Lindberg

17

and Marshall 1984, Brown and Haight 1992). Unlike crustaceans such as the Caribbean spiny

lobster, which aggregates in shelters with conspecifics (Childress and Herrnkind 1997), stone

crabs are solitary and share shelters only during mating periods (Wilber 1989). Menippe

mercenaria also dominate shelter access and outcompete other crustaceans such as P. argus

(Behringer and Hart 2017). Juvenile and young adult M. mercenaria inhabit the shallow hard-

bottom habitat of Florida Bay taking advantage of the multitude of solution holes and large

excavated loggerhead sponges as shelter. Larger individuals will emigrate to deeper sea grass

beds where they can excavate their own burrows as they outgrow the confines of hard-bottom

shelters.

Stone crabs support one of the most valuable commercial fisheries in the southeastern

United States and the Caribbean. The Florida fishery is the largest stone crab fishery in the

region, supporting an estimated $25 M annual commercial fishery. Monroe and Collier counties

lead the state in landings, producing more than half of the overall harvest (FWC 2017). Both

species of Menippe are managed as one fishery. The stone crab fishery is unique in that only the

claws are taken and crabs are released to regenerate new claws. This practice would seem to

make this fishery more sustainable than others, but studies have shown that the removal of claws

can cause high mortality, alter diet, and alter prey size selection (Duermit et al. 2015). The

fishery is managed differently in each state with some states allowing removal of both claws,

while others allow removal of only one. Florida allows the harvesting of both claws if they are

both of legal size. On average, male crabs will enter the fishery during age three and females

during age four (Gerhart and Bert 2008). If cyanobacteria blooms are affecting population size or

nutritional condition of M. mercenaria in Florida Bay, the largest regional contributor to this

valuable fishery could suffer.

18

I hypothesized that habitat degradation caused by persistent cyanobacteria blooms could

affect the nutritional condition of M. mercenaria by limiting prey availability or prey choice in

impacted regions of the Bay. A change in prey could also alter stone crab trophic position

causing further alterations throughout the food web. Shifts in prey availability or choice could

also affect site or shelter fidelity, ultimately causing crabs to emigrate from degraded regions to

more suitable habitat. Emigration or decreased nutritional condition could both have deleterious

effects on the stone crab fishery. Cyanobacteria blooms could also indirectly affect M.

mercenaria by killing sponges. If stone crabs use chemical cues exuded by hard-bottom

organisms, such as sponges, as a way of finding suitable habitat, navigating, or foraging, then the

degradation of hard-bottom could also have an ecological impact on behavior. I used three

indices to determine nutritional condition and employed stable isotope analysis of muscle tissue

samples to determine the trophic niche of M. mercenaria in degraded versus non-degraded

regions. A mark-recapture study was used to compare site and shelter fidelity between degraded

and non-degraded regions.

19

Figure 1-1. Map of Florida Bay (USA) lying between the mainland of Florida and the Florida

Keys. Photo courtesy of U.S. Geological Survey.

20

Figure 1-2. A large male Menippe mercenaria captured in Florida Bay in 2017. Photo courtesy

of author.

21

CHAPTER 2

THE EFFECTS OF HARD-BOTTOM HABITAT DEGRADATION ON NUTRITIONAL

CONDITION AND TROPHIC ECOLOGY OF MENIPPE MERCENARIA

Introduction

Habitat loss has been shown to have a major impact on species richness and biodiversity

in marine and terrestrial ecosystems (Ehrlich 1988, Dulvy et al. 2003). Not only can disturbances

to ecosystems affect organisms directly, but they can also cause changes in the community

structure. Anthropogenic stressors, such as nutrient input into coastal waters, have been shown to

alter ecosystem dynamics and condition such as productivity to higher trophic levels, low

dissolved oxygen content, and habitat loss in nearshore communities (Nixon et al. 1986, Short

and Burdick 1996, Breitburg et al. 1997). These alterations, among others, can result in drastic

changes to trophic structure as well as the condition of individual species.

When characterizing the condition of an organism, it is also important to take into

account prey that may be affected due to disturbances. For example, nutrient loading in Waquoit

Bay, Massachusetts has led to a decline in eelgrass habitat and an increase in macroalgae

biomass. This transition from eelgrass to macroalgae dominated communities has had further

implications on trophic structure and a decline in the abundance of bay scallop, Argopecten

irradians (Valiela et al. 1992). Effects such as these can have further implications on upper

trophic level organisms that rely on bay scallops as a food source, causing a bottom-up effect. In

another example, decreased nutritional condition and subsequent decline in Steller sea lion,

Eumetopias jubatus, populations in the Gulf of Alaska and off the Aleutian Islands was

attributed to a decline in the quality of prey available (Trites and Donnelly 2003). The condition

of a species may also be affected by habitat alteration or degradation. Nearshore habitat

alterations caused by coastal industrialization and chemical contaminants were shown to impact

the growth and condition of juvenile sole, Solea solea, in the English Channel. Sole grew slower

22

and had lower condition indices in areas with higher amounts of runoff and chemical

contaminants (Amara et al. 2007).

The loss of habitat due to cyanobacteria blooms in Florida Bay could be affecting M.

mercenaria directly or indirectly. Menippe mercenaria predominantly feed on hard-shelled

bivalves throughout their range (Menzel and Nichy 1958, Powell and Gunter 1968, Lindberg and

Marshall 1984, Brown and Haight 1992) with other potential prey in Florida Bay consisting of

gastropods and echinoderms (Pharo pers. obs.). Large areas of seagrass habitat in the Bay, often

adjacent to hard-bottom, can have > 75 taxa of gastropods and > 25 taxa of bivalves (Nizinski

2007) making these areas ideal foraging grounds. Species such as turban snails, hard clams, and

oysters can be plentiful in non-degraded hard-bottom regions of the Bay, but appear to be far less

common where blooms have drastically changed benthic trophic communities and bottom

structure (Pharo pers. obs.). Any potential change in prey abundance or diversity could

negatively affect the nutritional condition of M. mercenaria in degraded regions of Florida Bay.

A shift in prey or primary producers could also alter the trophic niche and nutritional condition

of M. mercenaria. Inter- and intra-species variability in lipid, protein, and fatty acid

concentrations have been shown to occur in several species of mollusks (Soriguer et al. 1997),

and such differences can then affect nutritional condition of predators such as M. mercenaria. I

compared nutritional condition and trophic position of M. mercenaria between degraded and

non-degraded regions of Florida Bay using several condition indices and muscle stable isotope

analysis.

23

Methods

Objective 1: To Determine the Effects of Hard-Bottom Habitat Degradation on the

Nutritional condition of M. mercenaria.

During the summer of 2017, crabs (n=123) were sampled from degraded and non-

degraded regions of Florida Bay by hand. Degraded regions of the Bay were chosen based on

previous literature and knowledge of non-degraded hard-bottom communities in the past. To

determine the effects of hard-bottom habitat degradation on the nutritional condition of M.

mercenaria, three indices were measured.

Immediately after removal from the water, 0.1 mL of hemolymph was extracted from the

most proximal leg sinus of the crab using a 27-g 1-mL tuberculin syringe and placed on an

industrial fluid refractometer to determine its refractive index (RI) to 0.5 units. The RI has been

used to assess nutritional condition in lobsters and other decapods since it is a reliable proxy for

serum protein (Musgrove 2001, Behringer and Butler 2006, Gutzler and Butler 2017); however,

serum protein levels of crabs are known to change with molt stage (Smith and Dall 1982) and

there is currently no method for assessing the molt stage of M. mercenaria.

The dry weight index (DWI) was also measured and compared between M. mercenaria in

degraded and non-degraded regions to indicate any changes in nutritional condition. This index

has been shown to be the preferred indicator for nutritional condition in crustaceans as it is not

affected by molt stage (Gutzler and Butler 2017). Crabs were iced for 10 min, euthanized and

dissected to obtain the hepatopancreas (Fig. 2-1). The hepatopancreas is an important organ in

decapods which stores lipids, carbohydrates and protein, provides digestive enzymes (Gibson

and Barker 1979), and is directly associated with nutritional condition (Rosemark et al. 1980).

Starvation studies in the western rock lobster, Panulirus longipes, have shown a stark decrease in

the mass of the hepatopancreas when starved (Dall 1974). The hepatopancreas of each crab was

24

weighed wet and then placed in a freeze drier for 72 h to obtain the dry weight. After dissection

of the hepatopancreas, the remaining carcass was placed in a drying oven for 24 h at 60° C to

determine its dry weight for use in an additional index. Hepatopancreas dry weight and crab wet

weight were used to determine the DWI calculated by Equation 2-1:

Dry Weight Index =Hepatopancreas Dry Weight

Animal Wet Weight × 100 (2-1)

The weight to carapace width (CW) ratio is another index that has been widely used in

nutritional condition studies of other decapods (Catacutan 2002, Oliver and MacDiarmid 2002).

CW was measured to 0.1 mm using Vernier calipers and dry weights were measured to 0.001 g.

The dry weight was the preferred choice to be used in this index as the wet weight of decapods

fluctuates with molt stage and water intake.

All statistical analyses were conducted using R software. Data used in the three indices

were found to be non-normal using a Shapiro-Wilks test. Transformations on the data were

attempted to achieve normality; however, data were not normal, therefore, indices were analyzed

using a non-parametric Kruskal-Wallis test.

Objective 2: To Determine the Effects of Hard-Bottom Habitat Degradation on the Trophic

Ecology of M. mercenaria

Stable isotope analysis is a widely used method in assessing trophic relationships within

communities (Peterson and Fry 1987, Wada et al. 1991, Post 2002). Stable isotopes of elements,

such as carbon and nitrogen, are used to determine the niche of different organisms and these

isotopes fractionate (relative change in abundance of the isotopes) when incorporated into

another organism. Isotopes are commonly referred to in terms of delta (δ) values which are the

measure of heavy and light isotopes. Delta values can be calculated by Equation 2-2:

δ X = [(Rsample / Rstandard)-1] x 103 (2-2)

25

where X is 13C or 15N and R is the ratio of 13C/12C or 15N/14N (Peterson and Fry 1987). Carbon is

typically used to indicate sources of primary production (Fry and Sherr 1984). δ13C values will

often be similar to the diet source of an individual, fractionating very little depending on the

protein content of prey (Fry and Sherr 1984, Peterson and Fry 1987). δ15N values indicate an

organism’s trophic level within the community and fractionate by approximately 3 ‰ per trophic

level (Minagawa and Wada 1984, Peterson and Fry 1987). The higher up in the food web an

organism is, the higher its δ15N value will be.

A subset of M. mercenaria (n=60) from the nutritional condition study (Obj. 1) were used

to compare the trophic position of crabs between degraded and non-degraded regions. Muscle

tissue samples were collected from the claws of crabs from degraded (n=30) and non-degraded

regions (n=30). Muscle tissue was used because of its slower turnover rate compared with other

tissue types, thus its stable isotope values can give an idea of long term (weeks to months)

trophic ecology (MacNeil et al. 2006, Madigan et al. 2012). Crabs were selected across similar

ranges of weight and CW to decrease variability between samples based on crab size. Tissue

samples were dried in a drying oven at 60°C for 24 h and then ground to a powder in a mill

(Wig-L-Bug; Crescent Dental). Samples were sent to the University of Florida Department of

Geological Sciences Stable Isotope Mass Spectrometry lab for stable isotope analysis. δ13C and

δ15N values of individual crabs were compared between degraded and non-degraded habitat to

test for differences. δ13C and δ15N values were also compared between sexes and size classes to

determine if either factor affected the carbon source or trophic position, respectively, of M.

mercenaria. All statistical analyses were conducted in R using a non-parametric Kruskal-Wallis

test.

26

Samples of turtle grass, Thalassia testudinum, red macroalgae, Laurencia sp., brown

branching sponge, Ircinia sp., and turban snails, Lithopoma tectum, were also collected from

hard-bottom habitat in degraded and non-degraded regions. Multiple composites were made

consisting of a range of 2-5 individuals each from these species sampled in degraded and non-

degraded habitat. δ13C and δ15N values were also obtained for these composites and a dual

isotope plot was constructed with M. mercenaria included to visualize the trophic niche of M.

mercenaria in relation to the community.

Results

Objective 1: To Determine the Effects of Hard-Bottom Habitat Degradation on the

Nutritional Condition of M. mercenaria

Crab size ranged from 34.8 to 101.7 mm CW and dry weight from 6.1 to 178.1 g. The sex

ratio of male to female was 1:1.46. Weight to CW ratios (χ2=0.577, df=1; p=0.448) (Fig. 2-2),

hemolymph refractive index (χ2=0.118, df=1; p=0.731) (Fig. 2-3), and DWI (χ2=0.641, df=1;

p=0.423) (Fig. 2-4) of M. mercenaria were not significantly different between degraded and non-

degraded regions. However, the DWI differed significantly between sexes (χ2=4.327, df=1;

p=0.038) (Fig. 2-5), with the DWI of females being significantly less compared to males. This is

most likely due to females utilizing energy and lipids from the hepatopancreas for reproduction.

To account for this difference, the DWI of each sex was compared separately between degraded

and non-degraded regions (Fig. 2-6). Females showed no significant difference in DWI in

degraded versus non-degraded habitats (χ2=0.136, df=1; p=0.713), although hepatopancreas

weight varied based on reproductive stage. No significant difference was found in DWI for

males between degraded and non-degraded habitats, indicating that nutritional condition was

similar (χ2=0.002, df=1; p=0.969).

27

Objective 2: To Determine the Effects of Hard-Bottom Habitat Degradation on the Trophic

Ecology of M. mercenaria

The CW of crabs selected for stable isotope analysis ranged from 34.8 to 97.8 mm and

the dry weight ranged from 6.1 to 147.1 g. The sex ratio of males to females was 1:1.5. The

weight to CW ratio of M. mercenaria used (n=30 in degraded and n=30 in non-degraded) did not

differ significantly between degraded and non-degraded regions (χ2=1.259, df=1; p=0.262) (Fig.

2-7). Stable isotope analysis revealed a significant difference in the trophic niche of M.

mercenaria between degraded and non-degraded regions (Fig. 2-8). The mean ( SD) δ13C

values of M. mercenaria in non-degraded and degraded regions were -12.78 0.86 and -9.76

1.30, respectively. The δ13C value of M. mercenaria in non-degraded regions was significantly

more negative than that of crabs in degraded regions (χ2=40.415, df=1; p<0.001) (Fig. 2-9A).

The mean ( SD) δ15N values of M. mercenaria in non-degraded and degraded regions were 6.78

0.34 and 6.72 0.53, respectively. δ15N values showed no significant difference between

degraded and non-degraded regions, but showed more variability in degraded regions (χ2=0.813,

df=1; p=0.367) (Fig. 2-9B). Comparisons were also made to distinguish whether trophic ecology

was related to sex (Fig. 2-10). Neither δ13C (χ2=0.018, df=1; p=0.892) (Fig. 2-11A) nor δ15N

(χ2=0.001, df=1; p=0.976) (Fig. 2-11B) values differed significantly between female and male M.

mercenaria.

Crabs were divided into two size classes, large and small, relative to the median carapace

width (72.6 mm) of the 60 samples taken. Comparisons were made between size classes to

determine whether size was correlated with δ15N or δ13C values (Fig. 2-12). Crabs within the

small size class ranged from 34.8 to 72.6 mm CW with a mean ( SD) of 59.4 9.94 mm. The

larger size class ranged from 72.6 to 97.8 mm CW with a mean ( SD) of 81.3 6.16 mm. δ13C

values did not differ significantly between small and large M. mercenaria (χ2=0.483, df=1;

28

p=0.487) (Fig. 2-13A). δ15N values also did not differ significantly between size classes

(χ2=3.526; df=1; p=0.06) (Fig. 2-13B); however, the nitrogen values were more variable among

smaller crabs.

Discussion

Habitat degradation in Florida Bay appeared to have little impact on the nutritional

condition of M. mercenaria, regardless of the metric used. However, stable isotope results

suggest a difference in the trophic niche of M. mercenaria between degraded and non-degraded

regions with an apparent shift in δ13C values. The δ15N values of M. mercenaria also appear to

be much more variable in degraded regions of the Bay.

Previous studies have reported a decline in species abundance and diversity after habitat

loss or degradation (Butler et al. 1995, Deegan et al. 2002, Hiddink et al. 2006), but few

compared species-specific nutritional condition between disturbed versus undisturbed

environments. Nutrient loading and chemical contaminants within sediments along industrialized

shorelines were shown to decrease growth rates and lower condition of juvenile sole, Solea

solea, in the English channel compared with shorelines with minimal anthropogenic impacts

(Amara et al. 2007). Ecosystem changes due to anthropogenic stressors have also been shown to

affect trophic structure and cause declines in abundance of important lower trophic level species

(Valiela et al. 1992). It appears the reoccurring cyanobacteria blooms in Florida Bay may also be

responsible for altering the trophic structure within degraded regions, but not necessarily

affecting the nutritional condition of M. mercenaria.

The similar nutritional condition of M. mercenaria between degraded and non-degraded

regions of the Bay suggests a resilience to this particular disturbance possibly due to a similarity

in prey availability between regions. Common prey items such as gastropods and bivalves

(Lindberg and Marshall 1984, Brown and Haight 1992) may be unaffected by cyanobacteria

29

blooms and thus just as abundant in degraded regions as in non-degraded regions. Previous

studies have shown M. mercenaria outcompete P. argus for shelter (Behringer and Hart 2017)

and degradation has been shown to affect lobster population densities (Butler et al. 1995), so

there is a possibility that prey availability would increase for M. mercenaria in the absence of

lobsters in degraded regions of the Bay.

If M. mercenaria do have access to similar prey within degraded and non-degraded

regions, the shift in δ13C values could indicate a potential change at a lower trophic level within

the community. The primary producer supporting the primary consumers or detritivores on

which M. mercenaria prey may have changed due to the chronic effect of reoccurring blooms,

and these changes are reflected bottom up through higher trophic levels. When multiple species

were plotted in a dual isotope plot (Fig. 2-14), it appeared that M. mercenaria in non-degraded

regions shared similar δ13C values to those of the macroalgae, Laurencia sp., suggesting this

species may be the primary producer indirectly supporting crabs through prey in non-degraded

regions. Laurencia sp. has been shown to be the main source of primary production supporting

hard-bottom communities (Behringer and Butler 2006) and occur as large mats in non-degraded

regions; however, these algal mats are far less apparent in degraded hard-bottom (Pharo pers.

obs.). M. mercenaria in degraded regions exhibited δ13C values intermediate to Laurencia sp.

and Thalassia testudinum, suggesting both primary producers may be supporting crabs indirectly

in degraded regions. It could be that M. mercenaria in degraded regions may be foraging more

heavily in adjacent seagrass beds when compared with crabs in non-degraded hard-bottom. Mere

geographical variability in δ13C values of sediment or primary producers could also contribute to

the observed differences.

30

Alternatively, M. mercenaria could be exhibiting a dietary shift if certain prey items are

negatively impacted by cyanobacteria blooms. Previous research has shown that molluscan

species, such as bay scallops and blue mussels, are negatively affected by the bloom forming

algae, Aureococcus anophagefferens, along the mid-Atlantic (Bricelj and Lonsdale 1997, Tracey

1988, Shumway 1990). In Florida, clearance rates of hard clams, oysters, and scallops have been

shown to be affected by the toxic bloom forming algae, Karenia brevis (Leverone et al. 2007). If

certain molluscan species are similarly affected by cyanobacterial blooms in Florida Bay, a shift

in diet to a more robust molluscan species could be occurring in bloom impacted regions.

Menippe mercenaria could also be exhibiting a shift in diet to more robust species such as sea

urchins or sea cucumbers, both of which crabs were observed to prey upon (Pharo pers. obs.).

The higher δ13C values seen in M. mercenaria in degraded regions of the Bay could be due to a

change in diet caused by the effects of the cyanobacteria blooms on species normally preyed

upon in regions not impacted by blooms. Furthermore, the variability observed in δ15N values of

M. mercenaria from degraded regions may be due to a more opportunistic or variable diet.

Variability in δ15N values of the same species have been attributed to diet variability among

individuals (Deniro and Epstein 1981). Due to possible effects of blooms on prey species, M.

mercenaria may be feeding on a variety of different organisms at different trophic levels

throughout degraded regions.

Despite extensive habitat degradation, M. mercenaria have been able to acclimate to this

disturbance and still maintain a similar condition. The most likely scenario being a more

opportunistic feeding strategy as seen by the results of stable isotope analysis, with M.

mercenaria in degraded regions utilizing adjacent seagrass habitat and feeding on a wider variety

of organisms. Further research should be focused on the impact of cyanobacterial blooms on

31

lower level trophic organisms in hard-bottom habitat, mainly prey species of M. mercenaria.

Examining the effects of cyanobacteria blooms on bivalves, gastropods, and echinoderms that M.

mercenaria would normally prey upon would give insight into other species that may be

impacted by the blooms. Collection of lower trophic level organisms and stable isotope analysis

could also be used to help further explain the alterations in trophic structure between degraded

and non-degraded regions. Furthermore, directly observing foraging habits of M. mercenaria

between degraded and non-degraded regions would yield valuable information on the effects of

cyanobacteria blooms on feeding behavior. Although algal blooms have been known to cause

decreased condition and mortality in many species (Paul et al. 2005, Leverone et al. 2007), the

ecological impacts of this particular cyanobacteria bloom do not seem to directly affect the

nutritional condition of M. mercenaria.

32

Figure 2-1. Underside of a stone crab carapace with hepatopancreas being removed for analysis.

Photo courtesy of author.

33

Figure 2-2. Box plots of the weight to carapace width ratios of M. mercenaria between degraded

and non-degraded regions. The dark shaded horizontal line indicates the median of

the data set. The box represents the middle 50% of scores for the data, while the

‘whiskers’ extending from the box represents the maximum and minimum values of

the data set. Individual points represent outliers.

34

Figure 2-3. Box plots of the hemolymph refractive index of M. mercenaria between degraded

and non-degraded regions. The dark shaded horizontal line indicates the median of

the data set. The box represents the middle 50% of scores for the data, while the

‘whiskers’ extending from the box represents the maximum and minimum values of

the data set. Individual points represent outliers.

35

Figure 2-4. Box plots of the dry weight index of M. mercenaria between degraded and non-

degraded regions. The dark shaded horizontal line indicates the median of the data

set. The box represents the middle 50% of scores for the data, while the ‘whiskers’

extending from the box represents the maximum and minimum values of the data set.

Individual points represent outliers.

36

Figure 2-5. Box plots of the dry weight index of M. mercenaria compared between males and

females. The dark shaded horizontal line indicates the median of the data set. The box

represents the middle 50% of scores for the data, while the ‘whiskers’ extending from

the box represents the maximum and minimum values of the data set. Individual

points represent outliers.

37

Figure 2-6. Box plots of the dry weight index of female and male M. mercenaria between

degraded and non-degraded regions. The dark shaded horizontal line indicates the median of the

data set. The box represents the middle 50% of scores for the data, while the ‘whiskers’

extending from the box represents the maximum and minimum values of the data set. Individual

points represent outliers.

38

Figure 2-7. Weight versus carapace width for M. mercenaria samples (n=60) for which muscle

tissue was analyzed for 13C and 15N values at degraded and non-degraded sites.

39

Figure 2-8. Dual isotope plot comparing δ13C and δ15N values of M. mercenaria between

degraded and non-degraded regions. Each point represents an individual crab. The

ellipses around both degraded and non-degraded points represent the 95% Confidence

Intervals of the data.

40

Figure 2-9. Box plots of the δ13C and δ15N values of M. mercenaria between degraded and non-

degraded regions. δ13C values are depicted on the left (A) and δ15N values on the right

(B). The dark shaded horizontal line indicates the median of the data set. The box

represents the middle 50% of scores for the data, while the ‘whiskers’ extending from

the box represents the maximum and minimum values of the data set. Individual

points represent outliers.

41

Figure 2-10. Dual isotope plot depicting δ13C and δ15N values between male and female M.

mercenaria. Each point represents an individual crab. The ellipses around both

degraded and non-degraded points represent the 95% Confidence Intervals of the

data.

42

Figure 2-11. Box plots of the δ13C and δ15N values of M. mercenaria between females and

males. δ13C values are depicted on the left (A) and δ15N values on the right (B). The

dark shaded horizontal line indicates the median of the data set. The box represents

the middle 50% of scores for the data, while the ‘whiskers’ extending from the box

represents the maximum and minimum values of the data set. Individual points

represent outliers.

43

Figure 2-12. Dual isotope plot depicting δ13C and δ15N values of M. mercenaria between size

classes. Each point represents an individual crab. The ellipses around both degraded

and non-degraded points represent the 95% Confidence Intervals of the data.

44

Figure 2-13. Box plots of the δ13C and δ15N values of M. mercenaria between size classes. δ13C

values are depicted on the left (A) and δ15N values on the right (B). The dark shaded

horizontal line indicates the median of the data set. The box represents the middle

50% of scores for the data, while the ‘whiskers’ extending from the box represents

the maximum and minimum values of the data set. Individual points represent

outliers.

45

Figure 2-14. Dual isotope plot comparing δ13C and δ15N values for multiple species between

degraded and non-degraded habitat. Ellipses represent 95% confidence intervals for

species. Each point for Laurencia sp., Lithopoma sp., and Thalassia testudinum

represent composite samples made of 5 individual samples each. Points shown for

Menippe mercenaria represent one individual crab each.

46

CHAPTER 3

THE EFFECTS OF HARD-BOTTOM HABITAT DEGRADATION ON SITE AND DEN

FIDELITY OF MENIPPE MERCENARIA IN FLORIDA BAY

Introduction

Movement patterns and fidelity to a particular habitat or shelter can have a significant

effect on the range and survival of a species. For example, these characteristics can affect the

ability of species to colonize new areas (Brousseau et al. 2002), survive harsh winter

temperatures (Wilber 1986), or escape hypoxic areas (Lenihan and Peterson 1998). Movement

can also be facilitated by habitat fragmentation (Micheli and Peterson 2001, Hovel and Lipcius

2001), food availability (Bearzi et al. 2006), and ability to find a mate (Diesel 1986). Factors

such as these can determine the extent to which a population can expand or contract. However, if

conditions are favorable in a particular habitat and resource availability is adequate, organisms

may show strong fidelity to areas. Adaptability also plays a large role in the structuring of

populations. The ability of an organism to utilize or adapt to multiple habitat types or conditions

can determine the dispersal of a population. Different marine organisms exhibit a variety of

different patterns of fidelity and movement.

Several species of fish have been shown to exhibit strong site fidelity to specific intertidal

rock pools and were also shown to return to original pools after being relocated 5-30 m away

(White and Brown 2013). The mangrove swimming crab, Thalamita crenata, uses visual cues

within its home range to relocate its shelter after foraging, showing high site fidelity (Cannicci et

al. 1995). Loggerhead turtles, Caretta caretta, do not necessarily show high fidelity to one area,

but are known for their homing abilities allowing them to locate the same foraging grounds and

nesting beaches repeatedly (Bowen 2004). P. argus has also been shown to have a homing

ability (Hernnkind and McLean 1971) and is able to forage and relocate its shelter afterwards or

use several shelters constantly moving back and forth among them (Bertelsen and Hornbeck

47

2009). In the northwest region of Florida, M. mercenaria was found to travel relatively short

distances between dens, exhibiting short term fidelity to particular inshore oyster reefs (Wilber

1986). Menippe mercenaria was also shown to exhibit low fidelity to single dens in a

manipulated shelter study. However, crabs were resighted within surrounding areas on

subsequent surveys (Lindberg et al. 1990). Prey depletion within the area was hypothesized as

the reason for the dispersal of crabs. Alternatively, the Asian shore crab, Hemigrapsus

sanguineus, shows very low site fidelity and high adult mobility making it a very successful

invader (Brousseau et al. 2002). Many of these examples of dispersal and homing abilities can be

attributed to visual cues, however, several species in the marine environment rely on olfaction to

locate suitable habitat.

In aquatic environments, olfaction plays a large role in the movement and behavioral

patterns of many species. Chemoreception is used for hunting, predator avoidance (Turner et al.

2003, Berger and Butler 2002, Behringer and Hart 2017), finding mates (Diaz and Thiel 2004,

Atema 1986, Hassler and Brockmann 2001), and disease avoidance (Behringer et al. 2018),

among other important roles. Crustaceans, such as the Caribbean spiny lobster use chemical cues

for many functions including avoidance of diseased conspecifics or to seek out healthy

conspecifics to shelter with (Anderson and Behringer 2013). Chemoreception is also used by

many species to cue settlement from their pelagic larval habitat (Forward et al. 2001, Raimondi

1988). For M. mercenaria, chemical cues from the brown algae Sargassum fluitans have been

shown to induce metamorphosis in juvenile M. mercenaria on the west coast of Florida

(Krimsky and Epifanio 2008). Considering the importance of chemoreception to many

crustaceans, including M. mercenaria, it would seem plausible that M. mercenaria would use

48

chemical cues exuded by benthic substrate as a means to navigate, find shelter, or find suitable

hard-bottom in which to reside.

The loss, degradation, or fragmentation of ecosystems could alter normal behavioral

patterns, leading to changes in movement or the reliability of chemosensory cues. Destruction of

important ecosystems has been shown to alter fish and crustacean communities and diversity

throughout many ecosystems (King 1998, Lenihan and Peterson 1998). Dramatic declines in the

abundance of seagrass-canopy-dwelling crustaceans were reported after massive seagrass die-

offs in Florida Bay due to algal blooms, however, benthic crustaceans increased in abundance

(Matheson et al. 1999). One of the many side effects of algal blooms are hypoxic or anoxic areas

(Paerl et al. 2001), which can kill or cause species to leave affected areas seeking more suitable

habitat. Off the coast of North Carolina, oyster reef habitat degradation in conjunction with

hypoxic or anoxic zones was shown to cause the blue crab, Callinectes sapidus, to completely

abandon their burrows in search of better habitat (Lenihan and Peterson 1998).

Within the hard-bottom habitat of Florida Bay, a wide variety of sponges cover the

benthos. A few species, such as Ircenia campana, are known as ‘stinker’ sponges and give off a

pungent odor when taken out of the water. These sponges saturate the water column with

chemicals that are most likely used by many marine organisms. Furthermore, juvenile and young

adult M. mercenaria are found residing in excavated loggerhead sponges (Fig. 3-1A) or solution

holes within the porous limestone sea floor (Fig. 3-1B) of the Bay. The loss of shelter could also

cause M. mercenaria to leave degraded areas in search of more suitable habitat. Presumably,

organisms will only inhabit areas with high prey availability. Prey diversity and abundance seem

to be relatively high with several species of bivalves and gastropods residing in the sediment or

on sponges and other structure (Pharo pers. obs.). However, prey density and diversity appear to

49

be much lower in impacted regions of the Bay, possibly due to the lack of habitat or due to the

direct effects of the cyanobacteria blooms. This potential difference in prey availability or

abundance could cause M. mercenaria to emigrate from degraded regions to locate more

desirable habitat with higher prey availability. The loss of these sponges, potential prey and

overall habitat due to persisting cyanobacterial blooms could be affecting fidelity and

chemosensory abilities of M. mercenaria. Here I investigated the effects of hard-bottom habitat

degradation in Florida Bay on fidelity and chemoreception of M. mercenaria.

Methods

Objective 1: To Determine the Effects of Hard-Bottom Habitat Degradation on the Site and

Den Fidelity of M. mercenaria.

During summer 2017, eight sites were selected in areas of Florida Bay impacted (Fig 3-

2A) and non-impacted (Fig 3-2B) by cyanobacteria blooms in recent years (e.g., 2007, 2013).

Three sites were identified in non-degraded areas and five in degraded areas. Areas were chosen

based on the presence and abundance of crabs to facilitate the study. Degraded sites were chosen

based on previous literature and the extent of the recent blooms. Sites were 30 m x 30 m plots

demarcated with weighted polypropylene line and cinderblocks. After plots were laid out, dens

within sites were located. Dens consisted primarily of crevices in or under loggerhead sponges

and solution holes in the porous limestone bottom. Only dens with M. mercenaria present on the

first survey day were used in the study.

Sites were surveyed by a team of three divers swimming back and forth between plot

borders within 1 m of each other to detect any shelters that were within the plots. Crabs were

collected by hand and lured from dens with the use of a sausage placed at the shelter entrance as

bait. Once captured, crabs were measured at the widest part of the carapace (CW), sexed, and

tagged while still under water. Tags consisted of American lobster claw bands color coded with

50

electrical tape to distinguish individual crabs from one another (Fig. 3-3A). Visible injuries and

shell disease (Rosen 1970, Iversen and Beardsley 1976) were also noted. After tagging, crabs

were placed back into their respective dens. Each occupied den was marked with a weight and

flagging tape with the corresponding color combination of the claw band written on the flagging

tape (Fig. 3-3B) to distinguish which crab belonged to which den. Sites and dens were then

surveyed again on days two, three, four and eight. On subsequent surveys, crabs were noted as

being present in their original den, present in another den within the plot, or absent from the plot.

New crabs that had moved into plots throughout the course of the eight days were also noted, but

not tagged.

A generalized linear mixed model (GLMM) in R program was used to analyze the

difference in fidelity to individual dens and sites between degraded and non-degraded sites. In

the model, ‘site’ was treated as the random effect to account for within site variance. Fidelity

between sexes was also analyzed given that male stone crabs are much more mobile than females

(Wilber 1989, Lindberg et al. 1990). A post hoc analysis of variance (ANOVA) was used to

determine any significant difference in fidelity between degraded and non-degraded sites.

A tag retention study was conducted in which 22 crabs were collected by hand from hard-

bottom habitat. Crabs were tagged as above and held in aquaria in the laboratory for 8 d. Crabs

were also fed various gastropods and bivalves throughout the course of the study. Tag retention

was recorded at the end of the 8 d period.

Objective 2: Response of M. mercenaria to Chemosensory Cues from the Loggerhead

Sponge, Spheciospongia vesparium.

Loggerhead sponges (Spheciospongia vesparium) are the largest sponge species in the

hard-bottom habitat of Florida Bay (Peterson et al. 2006) and are typically indicative of healthy,

non-degraded regions. Menippe mercenaria often utilize this species as shelter, excavating into

51

the sponge itself or occupying crevices below it. All sponges are filter feeders and pump water,

potentially releasing chemical compounds into the water column that could attract stone crabs.

Studies were conducted at the Keys Marine Laboratory in Layton, Florida. In the fall of

2016, loggerhead sponges ~30 cm in diameter were cut ~5 cm from the base of the sponge and

attached to 30 cm x 30 cm concrete pavers using zip-ties. Pavers with cut sponges were placed in

non-degraded hard-bottom areas to heal and attach to the paver over the next seven months (Fig.

3-4A). For the chemosensory experiment, two Y-mazes (Rebach 1996, Ratchford and Eggleston

1998, Diaz and Thiel 2004) were set up with two 100-L head tanks each (Fig. 3-4B). Y-mazes

(79 L) were constructed with 1.3 cm plywood and measured 94 cm long x 62 cm wide x 20 cm

tall with a central divide 72 cm long x 18 cm high (Fig. 3-4B). One head tank contained only sea

water to act as a control while the other contained the sponge treatment. The seawater used in the

trials was twice filtered to remove any large particulates. Shelters were constructed of 3 stacked

bricks (20 cm x 10 cm x 5 cm) and placed at the end of both chambers to attract crabs seeking

shelter.

Menippe mercenaria (n=29) were collected from hard-bottom habitat by hand and placed

in holding tanks prior to use in the experiment. Experiments were conducted at night to simulate

when crabs would be foraging and returning to shelter. For each trial, a sponge was placed in one

head tank for 30 min to allow chemical compounds being pumped from the sponge to diffuse

into the water. Head tank nozzles were then opened, allowing sponge treated and untreated water

to flow into either side of the choice chamber and into the center of the shelters at a rate of 5

ml/s. A dye test was used on each Y-maze to confirm that water and cues remained on their

respective sides and moved at a similar rate toward the drain at the base of the maze. Prior to

experimental trials, each crab was placed at the base of the maze behind a divider and allowed to

52

acclimate for 5 min. The divider was then lifted and the trial began. Crabs typically made a

choice of side after 30 min, after which final choice was recorded. A proportion test was used in

R to analyze the results.

Results

Objective 1: To Determine the Effects of Hard-Bottom Habitat Degradation on the Site and

Den Fidelity of M. mercenaria.

The tag retention study resulted in 86% retention over the course of 8 d, so it was

possible during the field study that some crabs lost their tags. However, based on retention rates

this was minimal and lost tags were often found outside the burrow of the crab that lost it. A total

of 106 crabs were tagged in the field, 40 in degraded sites and 66 in non-degraded sites. The CW

ranged from 30.7 to 93.1 mm in degraded and 50.4 to 94.7 mm in non-degraded sites. The sex

ratio of males to females was 1:3.1 within degraded and 1:1.2 in non-degraded sites. The GLMM

results indicated no significant difference in den fidelity between degraded and non-degraded

sites (χ2=0.829, df=1; p=0.396) (Fig. 3-5), nor was there a significant difference in site fidelity

among sites (χ2=0.844, df=1; p=0.996) (Fig. 3-6). When male and female crabs were analyzed

separately, no significant difference was found in female den fidelity (χ2=0.814, df=1; p=0.367)

(Fig. 3-7A), nor was there a significant difference in site fidelity (χ2=0.232, df=1; p=0.630) (Fig.

3-7B) between degraded and non-degraded sites. Similarly, no significant difference was found

in males for den fidelity (χ2=0.321, df=1; p=0.571) (Fig. 3-8A) or site fidelity (χ2=0.036, df=1;

p=0.849) (Fig. 3-8B) between degraded and non-degraded sites.

Objective 2: Response of M. mercenaria to Chemosensory Cues from the Loggerhead

Sponge, Spheciospongia vesparium.

A total of 28 Y-maze trials were performed. The CW of M. mercenaria ranged from 50.7

to 102.4 mm. Crabs chose the sponge treatment over seawater alone 67.9% of the time; however,

53

the difference between treatments was of borderline significance (χ2=3.571, df=1; p=0.058)

(Fig. 3-9).

Discussion

Den and site fidelity of M. mercenaria were found to be similar between degraded and

non-degraded sites, and there was no difference in fidelity between sexes either. Despite the

large loss in loggerhead sponges in Florida Bay, which are used as shelter by many organisms

(Pearse 1950, Butler et al. 1995, Behringer and Hart 2017), M. mercenaria were able to utilize

the abundance of solution holes within degraded sites. Chemosensory trials also indicated an

ability of M. mercenaria to detect chemical cues emitted by loggerhead sponges and the

subsequent attraction to these cues.

Habitat loss and degradation have been linked to population declines and decreased

species diversity in ecosystems globally (Brooks et al. 2002, Gray 1997, Valiela et al. 1992). For

example, results of many studies have shown stark declines in fish abundance and diversity on

coral reefs after bleaching events and subsequent loss of habitat for reef dwelling species (Bonin

2011, Pratchett et al. 2008, Pratchett et al. 2011). In the hard-bottom habitat of Florida Bay,

habitat degradation and the loss of sponges was shown to greatly reduce populations of P. argus

due to a decline in available shelters in degraded regions (Butler et al. 1995). However, results

reported here indicate habitat degradation in the Bay did not affect den or site fidelity of M.

mercenaria or its ability to find suitable shelter, most likely due to the availability of solution

holes and their ability to excavate burrows in seagrass.

Menippe mercenaria did not emigrate from degraded regions at a significantly faster rate

than those in non-degraded regions indicative of an acclimation to an altered habitat. However,

the results suggest that M. mercenaria spend more time in degraded regions than non-degraded

regions, albeit this result was not statistically significant. Presumably, the abundance of

54

resources, such as prey and shelter, within non-degraded habitat gives M. mercenaria the ability

to move around and forage in a larger area, resulting in limited fidelity to a particular site or

shelter. The lack of these resources in degraded regions could be forcing M. mercenaria to

forage more opportunistically, possibly taking advantage of adjacent seagrass habitat as foraging

grounds. Menippe mercenaria may also be feeding on a wider variety of organisms in degraded

regions, as shown by the variability in δ15N values of crabs in these regions, allowing them to

acclimate and remain in these regions. After massive seagrass die-offs in Florida Bay, also

associated with cyanobacteria blooms, benthic crustaceans were shown to increase in abundance

(Matheson et al. 1999), possibly moving in to capitalize on available space. A similar situation

could be occurring for M. mercenaria in affected regions of the Bay. Bottom trawling and

subsequent habitat disturbance and destruction has been shown to illicit the immigration of the

hermit crab, P. bernhardus, into recently disturbed areas to feed on injured or dead organisms

caused by trawling (Ramsay et al. 1996). This behavioral shift in foraging could also be

occurring in degraded regions, causing M. mercenaria to forage in adjacent areas or feed on

organisms injured by blooms. It could also be that the loss of sponges and other substrate has left

prey items much more vulnerable and available to M. mercenaria. Common prey items, such as

the gastropod, Lithopoma tectum, are found residing on many different sponges. The decline in

sponge biomass may have caused this species to forage on open bottom leaving them more

vulnerable to predation. Whether it is the relative similarity in prey availability or the shift in diet

and adaptability to an altered ecosystem, fidelity of M. mercenaria in degraded sites remained

similar to crabs in non-degraded sites. Menippe mercenaria may also be using chemical cues in

hard-bottom habitat as a means of relocating suitable habitat or shelter after foraging.

55

Chemoreception can play a large role in aquatic habitats and affect behavioral patterns of

many organisms including M. mercenaria (Behringer and Hart 2017, Krimsky and Epifanio

2008). The results of the Y-maze experiments, although of borderline significance, strongly

suggest that M. mercenaria may be attracted to chemical cues exuded by loggerhead sponges,

potentially as an indication of suitable habitat nearby. Substrate other than sponges, such as

corals and the red macroalgae (e.g., Laurencia sp.), could also be emitting chemical cues used by

M. mercenaria for navigation. Numerous organisms, such as the mangrove swimming crab,

Thalamita crenata, and Caribbean spiny lobster, Panulirus argus, use visual cues or an internal

map to locate shelters (Cannicci et al. 1995, Boles and Lohmann 2003). However, many species

rely heavily on chemoreception for navigation and homing (Croll 1983). The loss of sponges and

other substrate could be impacting movement patterns of M. mercenaria in degraded areas of

Florida Bay. Furthermore, chemical cues, such as those given off by the brown alga, Sargassum

fluitans, have been shown to elicit settlement and subsequent metamorphosis in M. mercenaria

on the west coast of Florida (Krimsky and Epifanio 2008) so the loss of habitat in Florida Bay

could be affecting settlement of larvae and population distribution.

Cyanobacterial blooms and the subsequent loss of hundreds of km2 of hard-bottom

habitat may affect the ecology of M. mercenaria in different ways. The fidelity shown by crabs

in degraded regions is comparable to non-degraded regions; however, the attraction that M.

mercenaria appears to show to the chemical cues emitted by loggerhead sponges may play an

important role in their ability to find suitable habitat, such that the loss of habitat could be

affecting chemoreception in M. mercenaria. Further research should investigate the general

movement, home range and foraging behavior of M. mercenaria in hard-bottom habitat as well

as their ability to relocate shelter after foraging via visual or chemical cues. General movement

56

of M. mercenaria could answer an important question regarding whether crabs are staying within

one large patch of hard-bottom habitat or if they are moving in between habitats. Furthermore,

the foraging behavior of M. mercenaria could give insight to where they chose to forage at night

and what habitats they utilize as foraging grounds in degraded versus non-degraded regions.

57

Figure 3-1. Common dens of M. mercenaria in hard-bottom habitat. Menippe mercenaria using a

loggerhead sponge as a den in non-degraded hard-bottom (A). Menippe mercenaria

using a solution hole as a den in a degraded region of Florida Bay (B). Photos

courtesy of author.

58

Figure 3-2. Degraded and non-degraded sites of Florida Bay used in fidelity study. Degraded

sites were located off Islamorada, FL in the impacted regions of blooms (A). Non-

degraded sites were located off Long Key and Marathon, FL in non-impacted regions

of blooms (B). Photos adapted from Google Earth.

59

Figure 3-3. Tags and weights used to mark individual crabs. Color coded American lobster claw

band used to tag largest claw on each crab (A). Stone crab tagged in a marked

loggerhead sponge den (B). Photos courtesy of author.

60

Figure 3-4. Sponges and Y-mazes used in chemosensory trials. Loggerhead sponge zip-tied to a

paver for use in chemosensory trials (A). Y-maze experiments with head tanks

containing seawater or sponge treated water (B). Photos courtesy of author.

61

Figure 3-5. Mean percentage of crabs recaptured within their original dens over the course of 8 d

in degraded and non-degraded sites. Error bars represent the 95% confidence

intervals.

62

Figure 3-6. Mean percentage of crabs recaptured on sites over the course of 8 d in degraded and

non-degraded sites. Error bars represent the 95% confidence intervals.

63

Figure 3-7. Fidelity of female crabs to individual dens and sites over the course of 8 d in

degraded and non-degraded sites. Mean percentage of female crabs recaptured within

their original dens (A) and on sites (B). Error bars represent the 95% confidence

intervals.

64

Figure 3-8. Fidelity of male crabs to individual dens and sites over the course of 8 d in degraded

and non-degraded sites. Mean percentage of male crabs recaptured within their

original dens (A) and on sites (B). Error bars represent the 95% confidence intervals.

65

Figure 3-9. Results of the final selections of M. mercenaria in chemosensory trials.

66

CHAPTER 4

CONCLUSIONS

Menippe mercenaria is an important and abundant benthic predator throughout its range.

In the Florida Keys, M. mercenaria supports higher trophic level organisms such as the red

drum, Sciaenops ocellatus, as well as a multimillion dollar commercial fishing industry (FWC

2017). Natural disturbances to species, such as M. mercenaria, could alter trophic structure

within hard-bottom communities, causing further trophic cascades as has been shown in other

species in the Florida Keys (Butler et al. 1995). Trophic cascades have been reported in many

ecosystems, marine and terrestrial, and can have deleterious effects, restructuring communities

and altering overall energy flow (Heck and Valentine 2006, Shurin et al. 2002, Wilmers et al.

2012). Therefore, it is imperative we understand the effects of persisting cyanobacteria blooms

on the ecology and biology of this ecologically and commercially valuable species.

The effects of cyanobacterial blooms in Florida Bay, although devastating to the benthos,

do not seem to negatively impact the condition of M. mercenaria. Nutritional condition index

comparisons between degraded and non-degraded regions of the Bay appear to be quite similar,

indicating no effect or an acclimation to this particular disturbance. If common prey items such

as bivalves and gastropods are also unaffected by blooms this could explain the similarity in

condition of M. mercenaria. No comparative studies between regimes have been done on the

abundance and condition of lower trophic level organisms within degraded regions of Florida

Bay, leaving this an open question.

Alternatively, M. mercenaria could be foraging more opportunistically in degraded

regions of the Bay possibly utilizing nearby seagrass beds as foraging grounds. This could be

explained by the apparent shift in trophic niche of M. mercenaria to a more positive δ13C value

in degraded regions similar to seagrass δ13C values. The variability in δ15N values could also be

67

attributed to M. mercenaria feeding on a number of different trophic levels within degraded

regions.

The loss of habitat also does not seem to affect the fidelity of M. mercenaria to a given

area. Although hundreds of km2 of sponge habitat has been lost due to reoccurring cyanobacteria

blooms, the abundance of solution holes within hard-bottom areas may provide adequate shelter

to M. mercenaria in these regions. However, the loss of sponges could be affecting crab’s ability

to find suitable habitat by limiting chemical cues emitted by substrate, such as sponges, in

degraded regions. The results suggest M. mercenaria is attracted to loggerhead sponges over sea

water alone, indicating an attraction towards healthy non-degraded hard-bottom. This could elicit

a problem, increasing time to find suitable habitat or shelter after foraging, leaving crabs

vulnerable to predation for longer periods of time. However, given that M. mercenaria show

short term fidelity within degraded regions of Florida Bay, it is possible that crabs are using

visual cues or an internal ‘map’ of their home range to locate their shelter (Cannicci et al. 1995)

as opposed to relying on chemical cues alone.

In this study, I aimed to discover the effects of habitat degradation due to cyanobacteria

blooms on the ecology and biology of the Florida stone crab in Florida Bay and although M.

mercenaria appear to exhibit a high resilience to this particular natural disturbance, further

studies are needed to address other questions. Several studies have shown the diet and prey

preference of M. mercenaria on the west coast of Florida (Menzel and Nichy 1958, Powell and

Gunter 1968, Lindberg and Marshall 1984, Brown and Haight 1992), but there is a lack of

knowledge on the ecology of stone crabs in the Florida Keys, the largest provider of stone crab

claws in the industry. Future research should be focused on potential prey of M. mercenaria in

Florida Bay and the effects of cyanobacteria blooms on these species. This information could

68

lead to a better understanding in the shift of trophic niche of M. mercenaria between degraded

and non-degraded regions. Although HABs have been shown to negatively impact important

marine species throughout the world (Bricelj et al. 1987, Butler et al. 1995, Gravinese et al.

2018), it would appear that the cyanobacteria blooms occurring in Florida Bay have little impact

on M. mercenaria, based on the metrics evaluated here.

69

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BIOGRAPHICAL SKETCH

Devon Pharo was born in Stuart, Florida in 1991. He grew up exploring everything

Florida had to offer from the beaches to the reefs in south Florida and the Florida Keys. His

family would often travel throughout the Caribbean, furthering his love of tropical marine

environments. He became enrolled at Santa Fe College in 2009 before transferring to the

University of Florida (UF) in 2011 to study Biology. Devon pursued his diving career at UF’s

dive program and became friends with a graduate student who would later introduce him to Dr.

Behringer. He volunteered in the Fisheries and Aquatic Sciences Department before graduating

in 2013 with a BS in biology and a minor in wildlife ecology and conservation. After graduating

he was employed at UF as a technician on several wildlife-related projects. In 2016, he was hired

by Dr. Behringer to work on an experiment examining the effects of Panulirus argus virus 1

(PAV1) on early benthic juvenile Caribbean spiny lobsters. He worked in the Florida Keys for 6

months on this project before applying to graduate school with Dr. Behringer’s lab. He was

accepted and immediately began working on a project examining the effects of hard-bottom

habitat restoration efforts in the Florida Keys. While working, Devon noticed the abundance of

young adult Florida stone crabs residing in hard-bottom communities within Florida Bay and

was intrigued by similar abundances of crabs in regions of the Bay degraded by reoccurring

cyanobacteria blooms. Hypotheses generated lead to the thesis research described in this

document.