Fire Ants on Sea Turtle Nesting Beaches in South Florida, Usa, and St

FIRE ANTS ON SEA TURTLE NESTING BEACHES IN SOUTH FLORIDA, USA, AND ST. CROIX, USVI

Danielle Kioshima Romais

A Thesis Submitted to the Faculty of

The Charles E. Schmidt College of Science

in Partial Fulfillment of the Requirements for the Degree of

Master of Science

Florida Atlantic University

Boca Raton, Florida

December 2013

ACKNOWLEDGMENTS

I would like to thank all who helped me, encouraged me, guided me and cheered for me throughout the development of this research. Thanks to my family, friends, co-workers, professors and mentors.

Dr. James K. Wetterer, I would like to thank you for the trust you invested in me from the very beginning. Thank you for the opportunity to conduct research in this topic. Thank you for the great ideas and the freedom to develop the work as my own. Dr. Jon Moore and Dr. Erik Noonburg, I would like to thank you for being members of this thesis committee. Thank you for the timely and valuable advice on methodology.

Thanks to Claudia Lombard from the U.S Fish and Wildlife Service in St.

Croix, USVI, for being so helpful during the Sandy Point survey. Thanks to Dr.

Mark Deyrup at Archibold Biological Station for conducting the final identification of all the ant species surveyed. Your expertise is unmatched. Thanks to Dr. Kirk

Rusenko and the Sea Turtle Research Team at Gumbo Limbo Nature Center for the logistic and GIS support during the surveys conducted in Boca Raton, FL.

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Thanks to the Environmental Science Program staff and faculty for setting the stage for learning and meaningful research to take place. Thanks to the Geosciences

Department for all the help in getting through this process as well. Thanks to the

Biology Department for the TA opportunity. Thanks to Dr. Gawlik and Dr. Markwith for allowing me to use their lab space during this research.

Thanks to Kavita Balkaran for conducting the survey in Jack Bay. Thanks to

André C. Kioshima for helping with survey in Pompano Beach. Thanks to Ariel

Shtauber for helping with survey in Boca Raton. Thanks to Dr. Sanford Porter for his expertise on fire ants and also for the images provided. Thanks to Dr. Julio C. de

Souza for images of our beautiful Brazilian Pantanal.

Thanks to Donna Selch, Mark Rochello for creating the sea turtle/fire ant interaction map. Thanks also to Hilton Cordoba for helping me design the maps for the study areas. You all rock!

Last, but certainly not least, I would like to thank my wonderful husband, Jeff

Romais, for the unconditional support and love during all this time. Thanks also to my beloved son, Kiyo, for being such an awesome trooper when I needed to be absent while working on this research. This is an accomplishment of our entire family, and I could not have done without the two of you!

ABSTRACT

Author: Danielle Kioshima Romais

Title: Fire Ants on Sea Turtle Nesting Beaches in South Florida, USA, and St. Croix, USVI.

Institution: Florida Atlantic University

Thesis Advisor: Dr. James K. Wetterer

Degree: Master of Science

Year: 2013

The red imported fire ant, Solenopsis invicta, is a South American native introduced in Alabama in the early 20th century. This predatory species has rapidly spread throughout the southeastern US and parts of the West Indies, inflicting great ecological and economic damage. For example, Solenopsis invicta is known to attack the eggs and hatchlings of ground nesting birds and reptiles. The ants swarm into the nests attacking hatchlings and diminishing their chance for survival. My thesis research aimed to survey the distribution of ants on sea turtle nesting beaches in South Florida and St.

Croix, USVI, and to evaluate the possible threat of Solenopsis invicta and other predatory ants to sea turtle hatchlings.

DEDICATION

I dedicate this work to my son, Kiyo Romais. He is my main source of inspiration, determination and strength. He is the reason I want to excel and make the world a little bit better.

Kiyo, I know that - when you grow up and become a man – you will make a difference in this world. Thank you for inspiring me ever since you were conceived to be a better mom, a better citizen, a role model to you. Thank you for you unconditional love and never ending curiosity. I love you, Kiyo, to the moon and back 1000 times!

FIRE ANTS ON SEA TURTLE NESTING BEACHES IN SOUTH FLORIDA, USA,

AND ST. CROIX, USVI

TABLES ...... i FIGURES ...... x 1. INTRODUCTION ...... 1 2. LITERATURE REVIEW ...... 5 2.1 Solenopsis sp...... 5 2.2 Fire Ants ...... 6 2.2.1 Native and introduced ranges ...... 7 2.2.2 Economic and health related impacts ...... 8 2.3 Fire ants as a threat for sea turtle hatchlings ...... 9 3 OBJECTIVES ...... 13 4 METHODS ...... 15 4.1 Study Areas ...... 15 4.1.1 Sandy Point Wildlife Refuge ...... 16 4.1.2 Jack Bay ...... 16 4.1.3 Boca Raton ...... 17 4.1.4 Pompano Beach ...... 19 4.2 Survey, Specimen Preservation and Identification ...... 21 4.2.1 Survey and Specimen Preservation ...... 22 4.2.2 Specimen Identification ...... 23 4.3 Sandy Point Wildlife Refuge Follow-Up Study: Phase I ...... 24 4.4 Baseline Distribution Survey: Phase II ...... 26 4.4.1 Pompano Beach, FL ...... 26 4.4.2 Jack Bay, St. Croix, USVI...... 27 vii

4.5 The Boca Raton Survey: Phase III ...... 28 4.6 Statistical Analysis ...... 29 5 RESULTS AND DISCUSSION ...... 32 5.1 Sandy Point Wildlife Refuge Follow-Up Study: Phase I ...... 32 5.2 Baseline Distribution Survey: Phase II ...... 35 5.2.1 Pompano Beach Survey ...... 35 5.2.2 Jack Bay, St Croix Survey ...... 39 5.3 The Boca Raton Survey: Phase III ...... 41 5.3.1 Baseline Survey Results ...... 41 5.3.2 Methodology Consistency Analysis ...... 44 5.3.3 Possible Threats to Sea Turtle Hatchlings ...... 53 5.4 Statistical Analysis ...... 56 6 FINAL REMARKS ...... 71

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TABLES

Table 1: Ant species surveyed in 2006 (Wetterer and Lombardo, 2010) ...... 25 Table 2: List of ant species present at Sandy Point, St. Croix, USVI ...... 33 Table 3: List of ant species in Pompano Beach, FL...... 36 Table 4: List of ant species found in Jack Bay, St. Croix, USVI...... 39 Table 5: List of ant species present in the Boca Raton study site – Initial Survey ...... 43 Table 6: List of species found in Boca Raton - Survey II ...... 46 Table 7: List of species found in Boca Raton - Survey III ...... 48 Table 8: List of species found in Boca Raton - Survey IV ...... 50 Table 9: Comparison per site between the four sampling efforts in Boca Raton, FL - Species observed based on abundance of individuals ...... 52 Table 10: Observed (O) versus Expected (E) sites with native and exotic species in 2006 and 2010 - Sandy Point, St. Croix, USVI (P>0.05) ...... 57 Table 11: Observed (O) and Expected (E) number os sites containing Solenopsis invicta and Solenopsis geminata in 2006 and 2010 - Sandy Point, St. Croix, USVI(P>0.05)...... 59 Table 12: Observed (O) and Expected (E) number of sites with native and exotic species in Sandy Point and Jack Bay (P<0.05) ...... 61 Table 13: Observed (O) and Expected (E) number of sites with native and exotic species in Pompano Beach and Boca Raton (P>0.05) ...... 63 Table 14: Observed (O) and Expected (E) number of sites with Solenopsis invicta in Pompano Beach and Boca Raton survey areas (P>0.05) ...... 65 Table 15: Observed (O) and Expected (E) number of sites with Solenopsis invicta in .... 67 Table 16: Observed (O) and Expected (E) number of sites with Solenopsis invicta in the Boca Raton survey area – four replicates (P>0.05) ...... 69

FIGURES

Figure 1: Solenopsis invicta (Image by S. Porter - USDA) ...... 7 Figure 2: Green turtle hatchling in Sandy Point, St. Croix, USVI ...... 10 Figure 3: Sandy Point, St. Croix Survey Area ...... 16 Figure 4: Jack Bay Survey Area ...... 17 Figure 5: Boca Raton Survey Area ...... 19 Figure 6: Pompano Beach Survey Area ...... 20 Figure 7: Bait-card placed along dune vegetation (Image by Kavita Balkaran) ...... 22 Figure 8: Point-Mount Method ...... 23 Figure 9: Identified Ant Species ...... 24 Figure 10: Ant Distribution in Sandy Point, St. Croix, USVI - Follow-up Survey. Green bars and (N) denotes native species, red bars are exotic species...... 34 Figure 11: Ant Distribution in Pompano Beach - Baseline Survey. Green bars and (N) denotes native species, red bars are exotic species...... 38 Figure 12: Ant Distribution in Jack Bay, St. Croix - Baseline Survey. Green bars and (N) denotes native species, red bars are exotic species...... 40 Figure 13: Ant Distribution in Boca Raton - Initial Survey. Green bars and (N) denotes native species, red bars are exotic species...... 44 Figure 14: Ant Distribution in Boca Raton - Survey II. Green bars and (N) denotes native species, red bars are exotic species...... 47 Figure 15: Ant Distribution in Boca Raton - Survey III. Green bars and (N) denotes native species, red bars are exotic species...... 49 Figure 16: Ant Distribution in Boca Raton - Survey IV. Green bars and (N) denotes native species, red bars are exotic species...... 51 Figure 17: Sea Turtle Nests and Fire Ant Presence in Boca Raton, Fl - 2012...... 55 Figure 18: Graph of frequency of native versus exotic species – the Sandy Point ant survey in 2006 against the Sandy Point ant survey in 2010 ...... 58 Figure 19: Graph of frequency of Solenopsis invicta and Solenopsis geminata per number of sites – the Sandy Point survey in 2006 against the Sandy Point survey in 2010 ...... 60 Figure 20: Graph of frequency of native and exotic species in Sandy Point and Jack Bay, St. Croix ...... 62

Figure 21: Graph of frequency of native and exotic species in Pompano Beach and Boca Raton ...... 64 Figure 22: Graph of frequency of Solenopsis invicta in Pompano Beach and Boca Raton, FL ...... 66 Figure 23: Graph of frequency of native and exotic species in Boca Raton ...... 68 Figure 24: Graph of frequency of Solenopsis invicta in Boca Raton - four replicates .... 70

1. INTRODUCTION

The world currently faces a biodiversity crisis where introduced species have population explosions while native species move toward extinction (Elton, 1958), which causes a significant shift in the dynamics of populations worldwide. With the advancement of technology and transportation, humans were able to colonize areas that would otherwise be uninhabitable. Anthropogenic changes to the environment seem to be a facilitating factor for exotic species establishment. Along with human settlements, a variety of species were introduced intentionally or accidentally. This does not imply that species were not able to move around prior to human intervention, but it correlates the speed at which these changes occur. As the movement of people increased both in intensity and in speed, the spread of introduced alien species have also increased

(Simberloff, 1997).

Biological invasion is a process in which a species introduced to a new and suitable habitat has a population explosion due to the lack of natural enemies and predators that could keep the population in check. This unbalanced advantage of the introduced species over the native ones can cause the latter to decline towards extinction.

Invasion by exotic species is the second greatest cause biodiversity loss in the world

(CBD, 1992). The first is the loss of habitat due to the conversion of natural areas for human use.

The impacts caused by invasive species may not be initially felt. Some species may not become a problem soon after introduction, and they may take years to establish a viable population. The time it takes for an invasion to occur does not make it a problem less than other environmental ailments because – if left unchecked and given the appropriate conditions – the problem can easily go from a manageable eradication effort to something that no longer can be controlled.

Natural habitats in South Florida are suitable for invasive species, including many exotic ant species that have been inadvertently introduced. These species are represented by a wide range of organisms from snails to reptiles, plants, mammals and insects. A comprehensive assessment conducted by Ferriter et al. (2008), listed 83 species of invertebrate organisms that is considered invasive in South Florida, six of which were ant species. Four species out of five invertebrates that were found in all the eight geographic regions studied were ants(Ferriter et al., 2008). Some of them – like the Burmese python

- have made headlines due to the negative impacts it is causing to the environment and the economy, as well as the possible threat to humans.

Biological invasions are directly or indirectly related to human sprawl and the environmental impacts caused by this phenomenon. In other words, sometimes the invasion takes place as a result of anthropogenic impacts, and the exotic species occupies an area after human disturbance has occurred. There are other times when the invasion

2 itself is the driving force to disturbance, causing massive impacts to the ecosystem

(Buczkowski, 2010).

Size does not matter when it comes to biological invasion. An example of this is the red imported fire ant (RIFA), Solenopsis invicta, considered to be an agricultural and urban pest in many introduced areas (Frank et al., 1997). This species is known for its powerful sting that can cause death to an adult human being in the most severe cases of allergic reaction. This species also presents economic impacts due to the costly attempts to eradicate the species in introduced areas. In ecological terms, the species exerts dominance over native ant populations and can successfully prey on vertebrates such as young bird and reptile hatchlings (Krahe, 2005). The increase or decrease of ant populations can also play a crucial role on the survival of species such as the sea turtles that nest on the South Florida beaches. Based on previous studies and field observations, the presence of predaceous exotic ants such as the RIFA along nesting beaches can impact the survival rate of sea turtle hatchlings nesting in South Florida and St. Croix

(Allen et al., 2001; Forys et al., 2005; Wetterer et al., 2007; J. K. Wetterer & Lombard,

2010).

One main goal with invasive species studies is to understand the invasion process and to devise efficient strategies to mitigate the impacts (Simberloff et al., 1997).

Determining whether or not an area has the potential to become invaded by an ant species requires a prior documentation of which ant species are present. Ant distribution surveys are regarded as fundamental in the development of a baseline understanding of which species are present in key areas for species conservation, diversity analysis and invasion. 3

In regards to the impacts caused by RIFA to sea turtles along the beaches in South

Florida and St. Croix, it is relevant to understand what species of ants are present in order to assess possible impacts caused by these species.

2. LITERATURE REVIEW

2.1 Solenopsis sp.

There are approximately 185 species worldwide that comprise the cosmopolitan genus Solenopsis (Hymenoptera: Formicidae) (Pitts et al., 2005). This genus belongs to the largest and most diverse subfamily of ants, Myrmicinae. Classification of the

Solenopsis genus is considered a difficult task by many taxonomists mainly because worker ants in most species are small and have few distinct characteristics useful in identification (Tschinkel, 2013).

Generally speaking, species belonging to this genus can be classified in two distinct groups: thief ants and fire ants. The first group is comprised of the majority of the species in the Solenopsis genus, characterized by the small and monomorphic workers.

Thief ants nest around other ants nests and “steal” the brood from the neighboring colonies, which is how they got their common name. The second group, the fire ants, is comprised of approximately twenty (20) species native to the New World, characterized by polymorphic workers and a powerful and painful sting (Pitts et al., 2005; Tschinkel,

2013).

The genus Solenopsis is usually associated with fire ants despite the fact that the majority of species in the genus are considered thief ants. This is because thief ants are generally smaller in size and more difficult to detect than the fire ants. Since they rarely come above ground, the thief ants are not collected as often as fire ants (Pacheco et al.,

2007). One example of thief ant is Solenopsis globularia, a species native in the

Caribbean.

The morphological characteristics of Solenopsis ant species are similar whether it is a thief ant or a fire ant. The difference lies primarily in the size of the workers, but even that can be sometimes misleading. Some fire ant specialists suggest that the one characteristic that distinguishes a fire ant from a thief ant within the Solenopsis genus is the individual’s ability to bite and sting, triggering the burning sensation associated with fire ants (S.Porter, personal communication, March 11, 2013).

2.2 Fire Ants

Only four of the twenty species of fire ants were identified as native to North

America. The other species are classified as part of a Neotropical assemblage known as the Solenopsis saevissima species-group (Pitts et al., 2005). Two of the sixteen species of

Neotropical fire ants (Solenopsis richterii and Solenopsis invicta) were inadvertently introduced to North America in the early 1900s.

Figure 1: Solenopsis invicta (Image by S. Porter - USDA)

Solenopsis invicta (Figure 1) is among the five species of ants listed as the worst biological invaders in the world (Sanders, 2011). Aside from health threats to human beings, S. invicta can also generate immense costs associated with efforts to control its spread (Lofgren, 1986). Although economic and health impacts are usually the primary reasons for controlling these invasive species, they can become major threats to the native wildlife and ant communities. In fact, invaders such as the red imported fire ant disrupt ecosystems to the point where native species communities are replaced by the exotic ones

(Mack et.al, 2000).

2.2.1 Native and introduced ranges

Fire ants were recognized to have come originally from South America by

Creighton and Löding in the late 1920s and early 1930s. The range of the two species of fire ants that invaded the United States (Solenopsis richterii and Solenopsis invicta) was further determined by Buren. Solenopsis invicta (RIFA) is native to the grasslands of

Southern Brazil and Northern Argentina, with its range centered along the Paraguay

River (Tschinkel, 2013). The species seems to have an ecological dominance over other ant species populations in its native range due to four main factors: number of individuals, recruitment system, aggressive and continuous foraging behaviors

(Calcaterra et al., 2008). The RIFAs are omnivorous and unicolonial (Parris et al., 2002;

Sanders, 2011). These characteristics along with a set of environmental factors such as habitat suitability, propagule pressure and lack of natural enemies in the introduced area increase their chance at becoming invasive.

The first introduction of the exotic fire ant, the black imported fire ant (Solenopsis richteri) happened in Mobile, Alabama around 1918. The introduction of the red imported fire ant (Solenopsis invicta) took place around the 1930s in Southern United

States (Pitts et al., 2005). The red imported fire ant spread throughout the southern portion of the country, out-competing its black exotic counterpart. Nowadays, the RIFA is found in approximately 1/3 of the United States, comprising most of the southern states and islands in the Caribbean. Its introduced range is considered by some experts as 4 to 5 times larger than the distribution in the native range.

2.2.2 Economic and health related impacts

Federal, state and municipal government agencies have spent millions of dollars to control the spread of S. invicta in the United States from the late 1950s to the early

1980s (Lofgren, 1986). It is estimated that if the spread of S. invicta is not monitored and controlled, the economic impact can range from $3 to $9 billion dollars for California alone in the next decade or so. These costs are associated with the application of 8 pesticides, property replacement or restoration, medical and veterinary treatments for sting reactions in humans, pets and livestock (Sanders, 2011).

The presence of S. invicta in urban areas is considered a nuisance to humans. The attacks can be by one worker’s multiple stings or by several individuals coordinating to sting simultaneously (Krahe, 2005). This results in an allergic response characterized by a burning sensation, excessive itching, the formation of a wheal and a blister-like pustule that can easily turn into a secondary infection (Lofgren, 1986). In some cases, the allergic reaction can be fatal due to the development of hypersensitivity reactions to the sting

(Lofgren, 1986; Sanders, 2011).

2.3 Fire ants as a threat for sea turtle hatchlings

There are seven identified species of sea turtle worldwide. Four of those seven species (Loggerhead, Leatherback, Green and Hawksbill) nest in South Florida and St.

Croix regularly (Figure 2). According to the International Union for Conservation of

Nature (IUCN) Red List, their status ranges from vulnerable to critically endangered.

This brings a sense of urgency to conservation efforts (2012).

Figure 2: Green turtle hatchling in Sandy Point, St. Croix, USVI

Sea turtles return to the beach in the vicinity of where they hatched to nest.

Nesting specificity makes them even more vulnerable to environmental changes.

Anthropogenic activities are considered a main driver for such changes, disturbing natural habitats and allowing the establishment of invasive species such as Solenopsis invicta. Sea turtle nests are often predated by a variety of organisms, both vertebrates and invertebrates.

Populations of S. invicta have been reported along the beaches of South Florida and many Caribbean islands, being found as far south as Key West (Davis et al., 2001;

Krahe, 2005; J. K. Wetterer & O'Hara, 2002; J. K. Wetterer & Lombard, 2010). These are commonly found along the area ranging from the dune into the wrack line (Allen et al.,

2001). Sea turtles nesting success depends on the area where the eggs are laid. Choosing to nest closer to the water line increases the risk of nest failure. The eggs might be washed away due to beach erosion processes during a storm surge. In order to guarantee a better chance at nest success, the sea turtles tend to nest closer to the dune line.

Therefore, the nests and hatchlings are more vulnerable to ant predations, particularly by exotic fire ants (Wetterer et al., 2007).

Fire ants can swarm a sea turtle nest soon after pipping and before emergence, which can take from several hours to several days (Wetterer et al., 2007). Some studies indicate that fire ant workers monitor the egg-chamber of freshwater turtles via underground foraging tunnels (Buhlmann & Coffman, 2001; Krahe, 2005). They can also act as scavengers feasting on the remains of hatchlings that have been attacked by other predators (Buhlmann & Coffman, 2001; Krahe, 2005). In either case, the chances for survival of the hatchlings attacked by the fire ants are greatly reduced. Even if the hatchling survives the initial attack, the exposure to a few fire ant stings can reduce its survival rate by 15% (Moulis, 1997). In her research Krahe (2005) indicated that for loggerhead turtles, the survival of hatchlings stung by RIFA decreased by over 50% ten days after hatching. There was no significant difference between green turtle hatchlings that have been stung versus those that were not stung by RIFA. Hatchlings that have been stung by fire ants show a more lethargic and paralyzed behavior. If they have been stung in the eye region, they display a “clouded-over eye” that may cause disorientation during crawling (Krahe, 2005).

3 OBJECTIVES

In order to mitigate the many problems caused by the spread of invasive alien species, it is important to determine their distribution and impact to native species. The dominance of invasive ants, in particular the red imported fire ants, can be first determined by ant population surveys that provide a baseline of data that can be used in future comparative analysis.

Seeing the impacts of invasive ants to endangered species such as sea turtles along shores worldwide, this study aimed to investigate the presence of exotic species of ants, mainly Solenopsis invicta, on four sea-turtle nesting beaches – two in South Florida and two in St. Croix, USVI. More specifically, our questions were:

1. Were there any significant changes in the overall ant distribution at Sandy Point,

St. Croix from 2006 (Wetterer & Lombard, 2010) to 2010 (present study)?

2. What species of ants are found in sea turtle nesting beaches in the surveyed areas

in South Florida and St. Croix, USVI?

3. What is the ratio between native and exotic ant species in these areas?

4. What is the percentage of sites with fire ants from genus Solenopsis found in these

areas?

5. Is the presence of surveyed species consistent if collected during multiple days?

As a way to answer these questions, this study aimed to document the current ant species distribution and predict ant dominance on sea turtle nesting beaches.

Documenting the distribution of ants is the first step to determine ant species dominance and invasion. Further studies can be developed based on baseline data provided in such distribution analysis.

The overall ratio between native and exotic ant species may infer the level of disturbance for the area. Generally, we expected to observe a higher number of invasive and/or exotic species in areas with more anthropogenic influence. The focus was on the distribution of fire ants from genus Solenopsis, mainly the red imported fire ants

(Solenopsis invicta) due to reported continuous attacks to sea turtle hatchlings and nests.

4 METHODS

In order to answer these questions, the present study was conducted in three different phases: Phase I – follow-up study in Sandy Point, St. Croix to observe any changes in species distribution for the area; Phase II – baseline surveys to document the current ant species distribution (exotic and native) on sea turtle nesting beaches in Jack’s

Bay, St. Croix and Pompano Beach, Florida; and Phase III – baseline ant distribution survey in Boca Raton, Florida aimed at investigating species present and analysis of consistency of the surveying methods conducted in multiple days.

4.1 Study Areas

The present study selected four areas, based on the high likelihood of being a sea turtle nesting beach. Two of the areas were located in St. Croix, USVI: Sandy Point

Wildlife Refuge and Jack’s Bay. The two other areas were located in South Florida: Boca

Raton and Pompano Beach. These areas have historically been selected by sea turtles as primary nesting grounds. Therefore, these areas are important for determining possible impacts of Solenopsis invicta on sea turtle hatchlings if in fact the invasive ant species is present in these sites.

4.1.1 Sandy Point Wildlife Refuge

Sandy Point Wildlife Refuge is a natural area monitored by the U.S. Fish and

Wildlife Service in St. Croix, USVI. The area is considered one of the most important sea turtle nesting areas in the island (Figure 3). It hosts the largest population of leatherback sea-turtles (Dermochelys coriacea) in the United States jurisdiction. Sandy Point is located at the southwest end of the island, south of Frederiksted. The area has a high touristic appeal, having an average of 11,000 visitors per year (USFWS, 2008).

Figure 3: Sandy Point, St. Croix Survey Area

4.1.2 Jack Bay

Jack Bay is part of two bays (Jack and Isaac Bays) located at the eastern end of St.

Croix. Comprised of white sandy beaches and upland forest, it is considered one of the 16 few pristine natural areas in St. Croix (Figure 4). The bays host the largest population of hawksbill and green sea-turtles in the island, and – due to their endangered status – these nesting areas are constantly monitored. The area is monitored by conservation efforts from The Nature Conservancy (TNC, 2013).

Figure 4: Jack Bay Survey Area

4.1.3 Boca Raton

Boca Raton is the southernmost town in Palm Beach County. The Boca Raton study area extends from the lookout Pavilion at the South Beach Park to the end of Red

Reef Park, along the beach. The beach in Boca Raton hosts sea-turtles of various species, but mainly loggerheads and green turtles nest in the area. The area is monitored by

Gumbo Limbo Nature Center in association with the Sea Turtle research conducted by

Florida Atlantic University. The starting point for the survey in this area was at life-guard tower 1 (26.34979 N and -80.06961 W) and extended to life-guard tower 12 (26.36684 N and -80.0682W) (Figure 5). The area is characterized by relatively dense dune vegetation with frequent foot trails for beach access. The beach is heavily used year-round by tourists and residents to the area.

Figure 5: Boca Raton Survey Area

4.1.4 Pompano Beach

Pompano Beach is located in central-east Broward County. The area is highly urbanized, and the beaches are usually busy with tourists and residents. The study area chosen in Pompano Beach is highly developed with little or no dune vegetation.

Buildings and other anthropogenic structures make up most of the shoreline in this area.

The area is also highly sought for tourism, fishing and recreational activities. There are numerous garbage cans and picnic areas that provide abundant food source for predator/scavenger ants such as Solenopsis invicta.

Figure 6: Pompano Beach Survey Area

Despite the disturbed conditions, sea turtles still come to this beach to nest.

Although efforts are made to protect nests, the heavy use of the beach and lack of dune vegetation to protect hatchlings from getting disoriented by street lights may be key at

20 impacting the survival of these already endangered species in the area. Adding to these factors, the presence of RIFAs in the area poses an even greater threat.

4.2 Survey, Specimen Preservation and Identification

Previous studies have indicated the expansion of Solenopsis invicta through South

Florida. This spread extends the species distribution as far south as the Florida Keys in the United States and in many of the Caribbean islands such as St. Croix (Davis et al.,

2001; Krahe, 2005; J. K. Wetterer & O'Hara, 2002; J. K. Wetterer & Lombard, 2010).

They are found along the coast as well as inland, being commonly associated with disturbed and urbanized areas. The red imported fire ants are considered a major threat to native ant communities as well as other organismal groups such as the sea turtles and shore birds nesting along the beach. The main question in this study is if there is any correlation between the presence of fire ants along the nesting beaches in South Florida and St. Croix and impacts on sea turtle hatchling survival.

The survey was conducted following the same format as described in Wetterer &

Lombard (2010). In general, surveys were conducted in the morning. A total of 12 – 15 baits were used per study area, with the exception of the survey conducted at Sandy Point

Wildlife Refuge – which was a more extensive area used as a follow-up for the study conducted in 2006 by Wetterer & Lombard. The surveys conducted in St. Croix used beach mile markers as guide to place the bait cards. In Boca Raton, cards were places on each life-guard tower that were located approximately 500 – 600 feet apart. In Pompano

Beach, the bait cards were placed in intervals of 50 – 100 meters since no permanent markers were observed at the surveyed site. 21

4.2.1 Survey and Specimen Preservation

Following the procedure described in similar studies (Wetterer et al., 2007;

Wetterer & O'Hara, 2002; Wetterer & Lombard, 2010), approximately 1 gram of water packed tuna bait was placed in a folded and properly marked index card. The cards were left in the sites for 2 hours, after which they were collected and individually placed into zip-lock bags (Figure 7). The bags were placed in a home freezer for approximately 2 hours in order to kill the ant specimens. The ants contained in each bag were visually inspected, described based on characteristics that were visible through analysis under a dissecting microscope and placed in vials of 95% ethanol for future analysis and preservation.

Figure 7: Bait-card placed along dune vegetation (Image by Kavita Balkaran)

4.2.2 Specimen Identification

A minimum of two individuals per site were mounted using the point-mount method (Figure 8), commonly used in preparing insects for long-term storage and later analysis. The individuals were selected based on their size (one larger and one smaller individual) and how complete was the specimen (Figure 9). For species that were represented by less than 10 individuals per bait, only one specimen was mounted. We used clear Elmer’s glue to mount the specimens as opposed to Hide glue which is commonly used. This product was chosen because it is easier to remove and it does not dry as fast as Hide glue.

Figure 8: Point-Mount Method

The mounted ants underwent three levels of identification. First, they were preliminarily identified by Danielle Romais, narrowing their identification based on genera. The second identification was done by Dr. James Wetterer, who identified most specimens by species. The third identification was conducted in order to either confirm or reject the two previous ones. It was done by Dr. Mark Deyrup, senior research biologist and ant specialist at the Archbold Biological Station in Venus, Central Florida. The

23 specimens were labeled according to standard methods of identification. The final storage of the specimens is yet to be determined.

Figure 9: Identified Ant Species

4.3 Sandy Point Wildlife Refuge Follow-Up Study: Phase I

In 2006, Wetterer and Lombardo (2010) conducted a survey at Sandy Point

Wildlife Refuge to investigate the prevalence of two species of fire ants (Solenopsis invicta and Solenopsis geminata) in this area. In this study, Wetterer and Lombardo

(2010) placed 33 baits along 1.7km stretch of beach at Sandy Point Wildlife Refuge in St.

Croix, USVI. In the survey conducted by Wetterer and Lombardo (2010), nine species of ants were identified. Five species were considered exotic and four were native. Fire ants

(Solenopsis invicta and Solenopsis geminata) occurred in 16 of the 33 baits, 48% of the sites. Solenopsis geminata was the most common fire ant found at Sandy Point during the

2006 survey, present in 33% of the baits. Solenopsis invicta was found in 15% of the sites. The native Dorymyrmex antillana was found in 14 of the 33 baits, comprising 42% of the baits (Table 1) (Wetterer and Lombardo, 2010). 24

Table 1: Ant species surveyed in 2006 (Wetterer and Lombardo, 2010)

Exotic Species Sites Native Species Sites

Solenopsis geminata 11 Dorymyrmex antillana 14

Solenopsis invicta 5 Brachymyrmex heeri 2

Paratrechina 4 Crematogaster steinheili 1

longicornis

Monomorium floricola 2 Monomorium ebenium 1

Paratrechina 1

bourbonica

In September of 2010, a follow-up distribution survey to the study conducted in

2006 (Wetterer and Lombardo, 2010) was conducted along the same stretch of beach at

Sandy Point Wildlife Refuge, St. Croix, USVI, as specified above. The starting point for the survey in this area was at marker 45 (17°41'3.50"N and 64°53'12.09"W), and it extended to marker 197 (17°41'0.83"N and 64°54'2.13"W). This survey took place in

September of 2010. The area was surveyed over a period of 3 days (09/18-09/20) due to storm weather conditions at the time. During the first two days, the survey was conducted in the morning, starting at 800 hours. In the last day, the survey was conducted in the afternoon, starting at 1630 hours.

The distribution of ants in the study areas was compared to the results obtained in the 2006 study conducted in the Virgin Islands. The analysis conducted was based on the different ant species found per bait-card. The species were divided into two categories: native and exotic. We expected to find at least the same number of exotic species as found by Wetterer and Lombardo during their study. Due to the high accessibility to this area, anthropogenic influence is evident, and it facilitates the spread and establishment of many exotic species such as Solenopsis invicta and Solenopsis geminata.

4.4 Baseline Distribution Survey: Phase II

Two baseline distribution surveys were conducted as part of this study in order to provide information about ant species present in these areas for future and more detailed studies. The areas in this initial survey were chosen because they are well documented sea turtle nesting sites. The two areas were: a stretch of beach in Pompano Beach, FL and in Jack Bay, St. Croix, USVI.

4.4.1 Pompano Beach, FL

There are no current records of ant surveys conducted in this area, so this study would provide a baseline data of ant species present along the Pompano Beach coast. Sea turtles nest on this beach, being more vulnerable to ant predation. This survey was conducted as a baseline analysis of the ant species present in the beach, so that future studies can be conducted as for possible impacts on sea turtle hatchlings. This baseline analysis aimed to investigate which ant species were found, and how their presence may impact the survival of endangered sea turtles.

The starting point for the survey in this area was located in alignment with 5th

Avenue and extended south to the beach access at Atlantic Boulevard (26°13'53.80"N and 80° 5'20.15"W). The survey area was divided by the Pompano Beach Fishing Pier.

The Pompano Beach survey was conducted in the morning of March 25th, 2012. A total of 15 baits were placed along the dune vegetation at an interval of 50-100 meters. The baits were placed at 800 hours in the morning and collected at 1000 hours, as specified above. The ant specimens were preserved, mounted and identified according to the methods explained above.

The area surveyed is highly influenced by anthropogenic activities. There are many picnic areas, playgrounds, garbage cans spread throughout the surveyed area. This area is also divided by the Pompano Beach Fishing Pier. Due to the high traffic of people and the use of the beach, it was expected a larger ratio of exotic versus native ants to be found in the survey.

4.4.2 Jack Bay, St. Croix, USVI.

The Jack Bay survey was conducted on December 18th, 2011 over a period of one day. A total of 15 bait cards were placed along the edge of the dune vegetation in the morning. The starting point for the survey in this area was at marker 6 (17°44'46.26"N and 64°34'41.60"W) and extended to marker 25 (17°44'49.18"N and 64°34'53.43"W).

The baits were placed at 800 hours in the morning at an interval of approximately 50-100 meters, including some sea turtle nests found in the area. The baits with specimens were collected at 1000 hours. The ant specimens were preserved, mounted and identified according to the methods explained above. 27

The Nature Conservancy did not report any sea-turtle nest in Jack Bay or Isaac

Bay being predated by fire ants – or other exotic ants. There are no records of ant distribution surveys conducted in this area of the island. Therefore, this survey serves as baseline data for any future ant distribution analysis conducted in Jack Bay, St. Croix.

Due to more pristine environmental conditions found in Jack Bay, St. Croix, a higher number of native ant species was expected.

4.5 The Boca Raton Survey: Phase III

The survey in Boca Raton was conducted to answer two questions in this research: first, we wanted to investigate which species were present in the area; and secondly, we wanted to repeat the survey in order to determine consistency in the methods used. The area was surveyed during 4 different times (06/12/2012; 06/16/2012;

06/29/2012; 07/06/2012) using the same methods described in Wetterer & Lombard

(2006) and other ant distribution studies (Wetterer et al., 2007; Wetterer & O’Hara,

2002).

Each bait card was placed by the lifeguard towers (1-12), starting at the South

Beach Park Pavilion and ending at the Red Reef Park. The GPS coordinates for each lifeguard tower was obtained, and a PVC stake marked the location where the bait card was first placed. This was to ensure that subsequent bait cards would be placed in the same location.

Each of the replicates was conducted in intervals ranging from 5 days to 15 days, depending on weather conditions. Ants are able to recall sites where they originally found

28 a food source due to chemical trails they leave as well as other cues they are able to gather from the environment (Collett et al., 2003). By spreading the collecting periods in longer intervals, we aimed to avoid dependence in the discovery and dominance of the bait cards which could skew our results. The shortest interval was 5 days between two survey efforts and the longest interval was 15 days.

The study area chosen in Boca Raton is the site where Gumbo Limbo Nature

Center conducts studies on sea turtle conservation, nesting and hatchling release. The dune and dune vegetation along the area is more protected and apparently less disturbed.

The area is highly sought for tourism, but local regulations seem to mitigate the impact.

Therefore, we expect to find a more diverse community of ant species. Exotic species of ants are also expected to be found in this area because of disturbance and other anthropogenic impacts that facilitate their establishment.

4.6 Statistical Analysis

The surveyed data was analyzed using the Chi-Squared Test in order to determine if the ratios between native species and exotic species found in a study area were significantly different from the each other. The critical alpha value to determine significance for this study was α = 0.05. The Chi-Squared Test (χ2) formula is given below. It computes the sum of the squared difference between the observed and expected values divided by the expected value.

∑

Equation 1: Chi-Square Formula

First, the native and exotic data collected at Sandy Point, St. Croix in 2006

(Wetterer & Lombard, 2010) was compared against the native and exotic data collected in 2010 – for this study. The ratio between Solenopsis invicta and Solenopsis geminata observed in the 2006 survey at Sandy Point was compared to the ratio of the same species observed in the 2010 survey of the same area. The ratio of native and exotic collected at

Sandy Point in the 2010 survey was compared against the ratio of native and exotic ant species collected in Jack Bay, St. Croix. Lastly, the ratio between native species versus exotic species observed in Pompano Beach was compared to the ratio between native and exotic species observed in Boca Raton – first survey replicate.

These analyses aimed to determine differences between surveying efforts in two distinct time periods as well as the difference in ratio among areas. The compared areas were paired based on geographic location. The areas in Florida, Pompano Beach and

Boca Raton, were compared to each other. The two areas in St. Croix, Sandy Point and

Jack Bay, were also compared to each other. The data obtained at Sandy Point in 2006 was compared to the same type of data obtained in 2010, determining temporal differences in the type of species found.

The survey replicates conducted in Boca Raton, FL were compared against each other to determine if the ratio between native and exotic species was significantly different. They were also analyzed in terms of the number of sites with RIFAs.

5 RESULTS AND DISCUSSION

5.1 Sandy Point Wildlife Refuge Follow-Up Study: Phase I

In a follow-up survey conducted in September of 2010, eleven different species of ants were found at Sandy Point Wildlife Refuge. Seven species were exotic, and four were native. Fire ants (Solenopsis invicta and Solenopsis geminata) were found in 38% of the baits. The native Dorymyrmex antillana was found in 25 of the 60 baits (41%), but the number of native species found was smaller than the number of exotic species.

Compared to the earlier survey, there was no significant difference on the number of sites with fire ants. Solenopsis geminata was found in more baits than Solenopsis invicta.

(Table 2).

Table 2: List of ant species present at Sandy Point, St. Croix, USVI Exotic Species Sites Native Species Sites

Solenopsis geminata 13 Dorymyrmex antillana 25

Solenopsis invicta 5 Solenopsis globularia 6

Nylanderia bourbonica 2 Monomorium ebenium 4

Tapinoma 2 Brachymyrmex heeri 1 melanocephalum

Monomorium floricola 1

Paratrechina longicornis 1

Tetramorium simillimum 1

One native species (Crematogaster steinheili) found during the 2006 survey was not found during the follow-up survey in 2010. The scientific name of one of the exotic species observed in 2006 changed from Paratrechina bourbonica to Nylanderia bourbonica (Lapolla et al., 2011). However, three different exotic species that had not been observed in 2006 were found in the 2010 survey. The overall number of native species was also lower than the number of exotic species. The number of native species found in both surveys was not different (4), but the species varied.

The native Dorymyrmex antillana showed dominance over the native species, being present in 25 of 60 baits. This may be an indicator that either the species is the native dominant in this area or that the presence of exotic ants has driven the number of

33 native ant species down. Although Dorymyrmex antillana was observed in the majority of baits, the number of individuals per bait was not high when compared to the two species of fire ants (Solenopsis invicta and Solenopsis geminata) (Figure 10).

Ant Distribution at Sandy Point, St. Croix, USVI

Solenopsis globularia (N) Monomorium ebenium (N) Monomorium floricola Brachymyrmex heeri (N) Nylanderia bourbonica

Dorymyrmex antillana(N) Paratrechina longicornis Tetramorium simillimum Solenopsis geminata Tapinoma melanocephalum Solenopsis invicta Solenopsis invicta Tapinoma melanocephalum

Species Found Species Solenopsis geminata Tetramorium simillimum Paratrechina longicornis Dorymyrmex antillana(N) Nylanderia bourbonica Brachymyrmex heeri (N) Monomorium floricola Monomorium ebenium (N) 0 5 10 15 20 25 30 Solenopsis globularia (N)

Number of Sites

Figure 10: Ant Distribution in Sandy Point, St. Croix, USVI - Follow-up Survey. Green

bars and (N) denotes native species, red bars are exotic species.

In baits where Solenopsis invicta was found (10 of the 60 baits), no other species were present. The other species of fire ants (Solenopsis geminata) was found in baits containing other ant species, but the density was visually greater for the fire ant species.

It is not possible to accurately infer that any impact is being caused by the presence of exotic ants, mainly the fire ant species, on sea turtle hatchlings and nests in

Sandy Point, St. Croix. Further studies analyzing the interaction between ants and sea turtles would have to be conducted in order to statistically make such affirmations.

However, there have been reports of ants swarming sea turtle nests and attacking hatchlings as they emerge (C. Lombard, personal communication, 2010). These reports along with the consistent presence of these exotic predaceous ants may be indicators of threat that the ants may impact the sea turtles.

5.2 Baseline Distribution Survey: Phase II

In order to understand which are the impacts caused by predator ants on endangered sea turtles, it is important to first know what species of ants are present in the area of interest. This information is obtained through a baseline study of the area. The presence of predatory ants, such as the RIFA, may pose an eminent threat to sea turtle hatchlings. Therefore, it is important to know if they are present in sea turtle nesting areas to be able to infer possible current or future threats. Baseline studies also serve as a predictor of future establishments, based on current range expansion trends.

5.2.1 Pompano Beach Survey

Humans have re-designed the shores at the city of Pompano Beach. Although the beach is still considered a desirable place for recreation, its function as a sea turtle nesting area is being constantly impacted by this new condition. Exotic ants such as the RIFA find this type of disturbed habitat suitable for their colonies because of the abundance in food sources available as a result of the increased number of trash and food residue left on the beach.

Since no previous ant distribution survey has been conducted in Pompano Beach, this survey aimed to serve as baseline for future studies. Due to the highly anthropogenic beach - with little or no dune vegetation and high recreational use - the number of exotic ant species was expected to be higher than the number of native species. The survey was conducted well within the sea turtle nesting/hatching season in order to provide us with the most accurate account of possible threat.

A total of three (3) species were identified in Pompano Beach, FL. One of the three was classified as native species. Two of the three were classified as exotic species

(Table 3).

Table 3: List of ant species in Pompano Beach, FL.

Exotic Species Sites Native Species Sites

Solenopsis invicta 5 Dorymyrmex bureni 6

Pheidole megacephala 1

Twelve (12) of the fifteen baits placed at the Pompano Beach Survey Area contained ants after the two-hour period. These twelve baits were classified as “active”.

Three baits had no ant specimens. These were classified “inactive”. One was removed by a beach goer and placed in a trash can nearby. The other two had no ants present.

Dorymyrmex bureni, a native species commonly known as “pyramid ant”, was found in six (6) of the twelve active baits, representing 50% of the total number of active

36 baits. The baits where these native species were present – and more predominant – were mainly located north of the Pompano Beach Fishing Pier. One bait at the end of the surveyed area had Dorymyrmex bureni present. Dorymyrmex bureni was the only native species found in this area during this baseline distribution survey (Figure 11). This species seems to be the only native that can be found in the same bait as the RIFA’s, suggesting that they may be sharing foraging territory (J.A.Moore, personal communication, November, 2013).

Pheidole megacephala, an exotic species commonly known as “big-headed ant”, was found in one (1) of the twelve active baits, representing 8% of the total number of active baits. This bait was located north of the Pompano Beach Fishing Pier, close to a picnic/recreational area. This exotic species is also considered invasive in many areas of the world, including Southern Florida. Pest control operators have reported that the bigheaded ant is becoming more pervasive as an invader because it displaces other ant species in the area (Warner & Scheffrahn, 2010).

Solenopsis invicta, the infamous red imported fire ant (RIFA), was present in five

(5) of the twelve active baits, representing 42% of the total number of active baits. These baits were predominantly located around and south of the Pompano Beach Fishing Pier.

As expected, the presence of RIFA was mostly in the portion of the surveyed area that suffered the biggest anthropogenic influence. The area is heavily used by tourists and local residents using the beach, picnic areas and playground adjacent to the beach. People also use the pier, and trash that is left lying around usually attracts these ants to the area

(Figure 11). 37

Ant Distribution in Pompano Beach

Solenopsis invicta

Pheidole megacephala Dorymyrmex bureni (N)

Pheidole megacephala SpeciesFound Solenopsis invicta

Dorymyrmex bureni (N)

0 1 2 3 4 5 6 7 Number of Sites

Figure 11: Ant Distribution in Pompano Beach - Baseline Survey. Green bars and (N)

denotes native species, red bars are exotic species.

Sea turtle nests in this area were not found very close to the vegetation line – where ants are usually found. RIFAs forage through tunnels that radiate from the mound.

Depending on the size of the territory, these foraging mounds can reach close to 100 meters from the main entrance to the mound. The presence of trash cans along the beach facilitate foraging in the areas of the beach closest to the sea turtle nests. Therefore, the presence of RIFAs in areas such as Pompano Beach may pose a significant threat to the survival of the hatchlings, considering that ants can attack them before or after they have emerged.

5.2.2 Jack Bay, St Croix Survey

Contrasting with Sandy Point, on the opposite side of St. Croix, Jack Bay is an area that is not heavily used by visitors. The area is monitored by The Nature

Conservancy because it is a nesting site for sea turtles. In conversation with the sea turtle specialist for TNC in St. Croix, she reported no knowledge of fire ant presence in Jack

Bay. Due to the overall pristine condition of the area, a higher number of sites with native species were expected.

Since there are no previous ant distribution survey for this area, this survey will serve as baseline data for future studies analyzing the possible threats of predaceous exotic ants on sea turtle hatchlings in the area. Twelve baits were placed along the vegetation at Jack Bay. All twelve sites were classified as active because ants were present in all baits.

A total of three (3) species of ants were identified in Jack Bay. Two of the three were classified as native species to Caribbean Islands. One species was classified as exotic (Table 4 ).

Table 4: List of ant species found in Jack Bay, St. Croix, USVI.

Exotic Species Sites Native Species Sites

Monomorium

floricola 1 Dorymyrmex antillana 10

Solenopsis globularia 1

As expected, the number of sites with native species was higher than those with exotic species of ants. In fact, eleven of the twelve baits contained native species of ants.

Only one bait contained exotic ant species. Dorymyrmex antillana, a native species of pyramid ant, was present in ten of the twelve baits. The species was present in 83% of the baits used for this study. Solenopsis globularia, a thief ant native to the Caribbean islands, was observed in one of the twelve baits – present in 8% of the total number of active baits (Figure 12).

Considering that Jack Bay is a more secluded area, with little and limited access to its shores, the presence of exotic species was expected to be very low. There was only one exotic ant species present in the baits used for this study in Jack Bay. Monomorium floricola, the exotic bicolored trailing ant, was present in one of the twelve baits. It represented 8% of the total number of active baits (Figure 12).

Ant Distribution in Jack Bay, St. Croix, USVI

Dorymyrmex antillana

(N)

Solenopsis globularia (N) Monomorium floricola

Solenopsis globularia (N) SpeciesFound Monomorium floricola Dorymyrmex antillana (N)

0 5 10 15 Number of Sites

Figure 12: Ant Distribution in Jack Bay, St. Croix - Baseline Survey. Green bars and (N)

denotes native species, red bars are exotic species. 40

5.3 The Boca Raton Survey: Phase III

The shores of Boca Raton, as it is true in much of the Florida coast, are considered a prime habitat for sea turtles to nest. As part of an effort to protect these already endangered species, Gumbo Limbo Nature Center houses a sea turtle hospital and research facilities that monitor the beach daily during nesting season. The presence of exotic predaceous ants was observed by the Sea Turtle Research Group that conducts nest surveys at the beach. However, no official ant distribution survey has been previously done in the Boca Raton beach area.

In order to provide sea turtle researchers with information about possible predators to hatchlings, this study carried out a preliminary ant distribution survey. The primary goal with this survey was to determine which ant species were present at the beach in Boca Raton, and – from the data obtained – help predict possible threats to sea turtles hatchlings. Secondly, the study analyzed whether the species collected would be consistent if surveys were repeated multiple times, and if it was possible to infer dominance of ant species based on distribution surveys. Four different surveys were conducted in the same sites. The intervals between each repeated survey ranged from 5 to

15 days.

5.3.1 Baseline Survey Results

Although Boca Raton is an area highly sought for tourism, the dune vegetation seems to be in healthy condition when compared to the study site in Pompano Beach. 41

Therefore, it was expected that the number of exotic species per site found would be lower or – at most – equal to the number of native species per site. Twelve bait cards used in this study were each placed by the lifeguard towers (1-12) located from South

Beach Park to Red Reef Park. This area is monitored by the Sea Turtle Research Team at the Gumbo Limbo Nature Center.

There have been informal observations of sea turtle nests being attacked by exotic predaceous ants. The Sea Turtle Research Team at Gumbo Limbo indicated that in such instances the ants would be classified as “fire ants”, but no further analysis of the ants was conducted. They also reported that ant mounds or trails would be treated with

Amdro®, specifically when ants were found near or on sea turtle nests.

A total of ten (10) species were observed on the bait cards for the first sampling of the area. Two of the ten species observed were classified as native to the area. The eight remaining species were classified as exotic (Table 5).

Table 5: List of ant species present in the Boca Raton study site – Initial Survey

Exotic Species Sites Native Species Sites

Solenopsis invicta 3 Dorymyrmex bureni 4

Tapinoma 2 Pheidole floridana 1 melanocephalum

Campanotus sexguttatus 1

Paratrechina 1 longicornis

Forelius pruinosus 1

Nylanderia bourbonica 1

Monomorium floricola 1

Tetramorium 1 bicarinatum

The number of native species was lower than expected despite the high usage of the beach. The native pyramid ant, Dorymyrmex bureni, was present in four of the twelve lifeguard tower sites. This species was found in 33% of the sites. Pheidole floridana, was classified as a native species to this area. It was present in one of the twelve sites, representing 8% of the total number of active baits (Figure 13). The two native species were found in a total of four (4) sites, making up 33% of the total number of sites surveyed.

Ant Distribution in Boca Raton - Survey I

Tetramorium bicarinatum Dorymyrmex bureni (N) Monomorium floricola Pheidole floridana (N) Nylanderia bourbonica Campanotus sexguttatus Forelius pruinosus Tapinoma melanocephalum Solenopsis invicta Paratrechina longicornis Paratrechina longicornis Solenopsis invicta SpeciesFound Tapinoma melanocephalum Campanotus sexguttatus Forelius pruinosus Pheidole floridana (N) Dorymyrmex bureni (N) Nylanderia bourbonica Monomorium floricola 0 1 2 3 4 5 Number of Sites Tetramorium bicarinatum

Figure 13: Ant Distribution in Boca Raton - Initial Survey. Green bars and (N) denotes

native species, red bars are exotic species.

There were eight ant species that were classified as exotic to this area. Solenopsis invicta was present in three (3) of the twelve sites on the first sampling. Tapinoma melanocephalum was present in two (2) of the twelve sites. The other exotic species were present in one of the twelve sites. The other exotic species were: Campanotus sexguttatus, Paratrechina longicornis, Forelius pruinosus, Nylanderia bourbonica,

Monomorium floricola and Tetramorium bicarinatum (Figure 13).

5.3.2 Methodology Consistency Analysis

In order to determine if the species present at a surveyed site accurately represent the distribution of ant species at sea turtle nesting beaches, this study conducted four

44 distinct and independent surveys at the Boca Raton study area. The four different surveys were conducted on intervals ranging from 5 to 15 days, depending on weather conditions.

Time of day and placement of bait cards were kept constant throughout the different sampling efforts.

Although variations on species found at each site were observed from one sampling to the other, overall the species were consistently found in the same sites. It was observed that the species that did vary were the ones found in least numbers per bait.

These species were not considered “dominant” per each site.

As stated earlier, the surveying efforts in Boca Raton were repeated four times in order to analyze consistency of results. The initial survey served as a baseline for ant distribution in this study area. The results for the initial survey are displayed and discussed in the previous section of this document.

On the second survey, a total of eleven (11) species of ants were observed. Three

(3) of the eleven species were classified as native to the area. The remaining eight (8) species were classified as being exotic.

Table 6: List of species found in Boca Raton - Survey II

Exotic Species Sites Native Species Sites

Campanotus Dorymyrmex 2 2 sexguttatus bureni

Technomyrmex Campanotus 2 2 difficilis floridana

Forelius pruinosus 2 Pheidole floridana 2

Nylanderia bourbonica 2

Solenopsis invicta 1

Cardiacondyla emeryi 1

Tetramorium 1 bicarinatum

Tetramorium 1 simillimum

Ant Distribution in Boca Raton - Survey II

Tetramorium simillimum Dorymyrmex bureni (N) Tetramorium bicarinatum Pheidole floridana (N) Nylanderia bourbonica Campanotus floridana (N) Forelius pruinosus Cardiacondyla emeryi Campanotus sexguttatus Technomyrmex difficilis Solenopsis invicta

Solenopsis invicta Technomyrmex difficilis SpeciesFound Campanotus sexguttatus Cardiacondyla emeryi Campanotus floridana (N) Forelius pruinosus Pheidole floridana (N) Nylanderia bourbonica Dorymyrmex bureni (N) Tetramorium bicarinatum 0 0.5 1 1.5 2 2.5 Tetramorium simillimum Number of Sites

Figure 14: Ant Distribution in Boca Raton - Survey II. Green bars and (N)

denotes native species, red bars are exotic species.

Solenopsis invicta was present in one site during this survey. There were four exotic species present in the greatest number of sites (2). These species were:

Campanotus sexguttatus, Technomyrmex difficilis, Forelius pruinosus and Nylanderia bourbonica (Figure 14). One of the bait cards (site number 5) was missing at the time of collecting. Logistic issues at the time of collecting the bait cards may have skewed the results for this survey. The bait cards were left in site longer than the desired two hours due to delay on getting to each site.

Table 7: List of species found in Boca Raton - Survey III

Exotic Species Sites Native Species Sites

Solenopsis invicta 4 Dorymyrmex bureni 5

Nylanderia bourbonica 2 Pheidole floridana 1

Technomyrmex difficilis 1

Tetramorium simillimum 1

Forelius pruinosus 1

Campanotus sexguttatus 1

Monomorium floricola 1

Tetramorium

bicarinatum 1

On the third survey, a total of 10 species were observed. Two of the ten species were classified as native while eight were classified as exotic (Table 7). Five sites out of twelve surveyed (42%) contained native species. Nine sites contained exotic species

(75%). Many species were found in combination with others in one single site.

Solenopsis invicta was present in four (4) out of twelve sites (33% of total sites).

Dorymyrmex bureni was present in five (5) of the twelve sites (42%) (Figure 15).

Ant Distribution in Boca Raton - Survey III

Tetramorium bicarinatum

Monomorium floricola Dorymyrmex bureni (N) Nylanderia bourbonica Pheidole floridana (N)

Forelius pruinosus Campanotus sexguttatus Tetramorium simillimum Solenopsis invicta

Technomyrmex difficilis Technomyrmex difficilis

SpeciesFound Solenopsis invicta Tetramorium simillimum Forelius pruinosus Campanotus sexguttatus Nylanderia bourbonica Pheidole floridana (N) Monomorium floricola Dorymyrmex bureni (N) Tetramorium bicarinatum 0 1 2 3 4 5 6 Number of Sites

Figure 15: Ant Distribution in Boca Raton - Survey III. Green bars and (N) denotes

native species, red bars are exotic species.

On the fourth survey, a total of eleven (11) species were observed. Two of the eleven species were classified as native to the area. Nine species were classified as exotic

(Table 8).

Table 8: List of species found in Boca Raton - Survey IV

Exotic Species Sites Native Species Sites

Solenopsis invicta 3 Dorymyrmex bureni 6

Nylanderia bourbonica 3 Pheidole floridana 1

Tetramorium simillimum 2

Technomyrmex difficilis 1

Campanotus sexguttatus 1

Paratrechina longicornis 1

Monomorium floricola 1

Wasmania auropunctata 1

Tetramorium bicarinatum 1

Native species were present in seven of the twelve sites. Exotic species were present in ten of the twelve sites. Many species (exotic and native) were found in combination with other species per site (Figure 16).

Ant Distribution in Boca Raton - Survey IV

Tetramorium bicarinatum Dorymyrmex bureni (N) Wasmania auropunctata Pheidole floridana (N) Monomorium floricola Nylanderia bourbonica Campanotus sexguttatus Tetramorium simillimum Solenopsis invicta Technomyrmex difficilis Paratrechina longicornis Paratrechina longicornis

SpeciesFound Solenopsis invicta Technomyrmex difficilis Campanotus sexguttatus Tetramorium simillimum Pheidole floridana (N) Dorymyrmex bureni (N) Nylanderia bourbonica Monomorium floricola 0 1 2 3 4 5 6 7 Number of Sites Wasmania auropunctata

Figure 16: Ant Distribution in Boca Raton - Survey IV. Green bars and (N) denotes

native species, red bars are exotic species.

The species found in greatest abundance (number of individuals greater than 100) when bait was shared with other species were classified as “dominant” over the other species. In some cases, the bait cards contained only one species of ant. These species were considered “dominant” as well (Table 9). “N/A” indicates that the bait was either missing or with no ants present.

Table 9: Comparison per site between the four sampling efforts in Boca Raton, FL -

Species observed based on abundance of individuals

Exotic Survey 1 Survey 2 Survey 3 Survey 4

Solenopsis 3 1 4 3 invicta Nylanderia 1 2 2 3 bourbonica Tetramorium 1 1 2 simillimum Tetramorium 1 1 1 1 bicarinatum Campanotus 1 2 1 1 sexguttatus Technomyrmex 2 1 1 difficilis Forelius 1 2 1 pruinosus Monomorium 1 1 1 floricola Paratrechina 1 1 longicornis Tapinoma 2 melanocephalum Wasmania 1 auropunctata Cardiacondyla 1 emeryi Native Survey 1 Survey 2 Survey 3 Survey 4

Dorymyrmex 4 2 5 6 bureni Pheidole 1 2 1 1 floridana Campanotus 2 floridana

Overall, the species observed at each bait card are consistently found in the sites.

There are some general variations in the results obtained, which was expected due to weather condition and time that the bait was left at each site. Although it may not be possible to determine dominance, it is reasonable to assume that these species are currently distributed in this area. Therefore, based on these results, the bait method used to survey ants in this study has shown consistent results in the species observed. Its aim was to survey for dominant ant species in the area.

5.3.3 Possible Threats to Sea Turtle Hatchlings

Several studies have indicated that RIFAs are a threat to ground nesting birds and reptiles (Greenwood, Palmer, & Richardson, 2010; Parris et al., 2002; Wetterer et al.,

2007; Wetterer & Hara, 2002). There have been reports about fire ant attacks on sea turtle hatchlings. These attacks happen either at the time of emergence or soon as the turtles have pipped. Ants can also monitor the egg chamber as a way of “knowing” when it is appropriate to swarm the nest. Fire ants can forage as far as mid-beach, so nests far from the vegetation can still be affected by ant attacks (Krahe, 2005; Wetterer et al., 2007).

In 2012, the same year when the ant distribution survey was conducted for this study, Gumbo Limbo Nature Center reported a total of 1143 nests along the entire stretch of beach monitored by their Sea Turtle Research Team (Rusenko et al., 2012). The presence of RIFAs poses a significant threat to the survival ability of sea turtle hatchlings because they are most vulnerable to predators at that stage.

This study was able to determine which sites were most likely to have RIFAs.

Solenopsis invicta was observed in five of the twelve sites in Boca Raton. This data was geo-referenced in ArcGIS and combined with the data obtained by Gumbo Limbo Nature

Center (Rusenko et al., 2012) on sea turtle nests for the 2012 nesting season. Considering that sea turtle hatchlings are potential targets for RIFAs attacks, the presence of

Solenopsis invicta (RIFAs) can be considered a threat to sea turtle nesting and hatchling survival and success (Figure 17).

Figure 17: Sea Turtle Nests and Fire Ant Presence in Boca Raton, Fl - 2012

Solenopsis invicta can forage very large areas, depending on colony size. The foraging territory can be adjusted based on food availability, but it may also be strongly related to the number of workers and territory size. Studies have indicated that colony ranges from 12 to 197 m2 (Dhami, 2008). Based on this information and the data obtained on sea turtle nests, fire ants may pose significant threat to hatchlings if population density of the invasive species is not kept under control.

5.4 Statistical Analysis

When comparing the number of sites with native and exotic species surveyed in

2006 against those surveyed in 2010 at Sandy Point, St. Croix, USVI, the p-value obtained using the chi-squared formula was 0.26, which is greater than the critical alpha value of 0.05. This indicates that the ratio of native and exotic species was not significantly different at Sandy Point from the 2006 survey to the 2010 survey. The table below indicates the observed values and the expected values used in this analysis (Table

10). The bar graph was used to indicate the frequency of native and exotic species during each of the surveyed periods (Figure 18).

Table 10: Observed (O) versus Expected (E) sites with native and exotic species in 2006

and 2010 - Sandy Point, St. Croix, USVI (P>0.05)

Observed

Natives Exotics Total Surveyed Sites

2006 18 23 33

2010 30 31 60

Grand Total 48 54 93

Expected

Year/Sites Natives Exotics Total Surveyed Sites

2006 17.03 19.16 33

2010 30.97 34.84 60

Grand Total 48 54 93

p-value

0.26

Frequency of Native and Exotic Species per number of sites - Sandy Point Survey 2006 vs. 2010

Exotics

2010 Species Natives 2006

0 5 10 15 20 25 30 35 Number of Sites

Figure 18: Graph of frequency of native versus exotic species – the Sandy Point ant

survey in 2006 against the Sandy Point ant survey in 2010

Since Solenopsis invicta and Solenopsis geminata are the two species of fire ants present in St. Croix, this study investigated the frequency that they were observed. The survey conducted in 2006 observed Solenopsis geminata in 11 sites, and Solenopsis invicta was observed in 5 sites. In 2010, Solenopsis geminata was detected in 13 sites while Solenopsis invicta was still detected in 5 sites in Sandy Point, St. Croix. The ratio of sites with Solenopsis invicta and Solenopsis geminata surveyed at Sandy Point in 2006 did not significantly differ from the 2010 survey. The p-value obtained from the chi- square test was 0.82, which is greater than 0.05. This indicates that the number of sites containing Solenopsis invicta and Solenopsis geminata at Sandy Point did not vary significantly from the 2006 survey to the 2010 survey. The table below indicates the observed values and the expected values used in this analysis (Table 11). The bar graph

58 shows the frequency of Solenopsis invicta and Solenopsis geminata at Sandy Point during the 2006 and the 2010 surveys (Figure 19).

Table 11: Observed (O) and Expected (E) number os sites containing Solenopsis invicta

and Solenopsis geminata in 2006 and 2010 - Sandy Point, St. Croix, USVI(P>0.05)

Observed

S. geminata S. invicta Total Surveyed

Sites

2006 11 5 16

2010 13 5 18

Grand Total 24 10 34

Expected

S. geminata S. invicta Total Surveyed

Sites

2006 11.29 4.71 16

2010 12.71 5.29 18

Grand Total 24 10 34

p-value

0.82

Frequency of Solenopsis invicta and Solenopsis geminata - Sandy Point Survey 2006 vs. 2010

S. invicta

2010 Species S. geminata 2006

0 5 10 15 Number of Sites

Figure 19: Graph of frequency of Solenopsis invicta and Solenopsis geminata per number

of sites – the Sandy Point survey in 2006 against the Sandy Point survey in 2010

The two sites surveyed in St. Croix for this study are remarkably different from each other. While Sandy Point is sought for tourism, Jack Bay is more secluded. The present study investigated if this difference could be detected in the ant distribution of each area. The results of the chi-squared test indicated that the ratio of native species versus exotic species at Sandy Point was significantly different than that of Jack Bay, St.

Croix. The p-value was 0.007, which is less than the alpha critical value of 0.05 (Table

12). The bar graph indicates the frequency at which native and exotic ant species were observed in Jack Bay versus Sandy Point (Figure 20). The two species of fire ants,

Solenopsis invicta and Solenopsis geminata, were not detected in Jack Bay. This can possibly be related to the environmental conditions of the area, but further study is needed in order to investigate this matter.

Table 12: Observed (O) and Expected (E) number of sites with native and exotic species

in Sandy Point and Jack Bay (P<0.05)

Observed

Natives Exotics Total Surveyed

Sites

Sandy Point 30 31 60

Jack Bay 11 1 12

Grand 41 32 72

Total

Expected

Natives Exotics Total Surveyed

Sites

Sandy Point 34.17 26.67 60

Jack Bay 6.83 5.33 12

Grand Total 41 32 72

p-value

0.007

Frequency of Native and Exotic Species per number of sites - Sandy Point vs. Jack Bay, St. Croix, USVI

Exotics

Jack Bay Species Natives Sandy Point

0 10 20 30 40 Number of Sites

Figure 20: Graph of frequency of native and exotic species in Sandy Point and Jack Bay,

St. Croix

The same comparative analysis was conducted for the two study areas in South

Florida – Pompano Beach and Boca Raton. A slight difference was expected to be observed between the two areas because Pompano Beach appears to be more impacted by anthropogenic use than Boca Raton. The surveyed area in Pompano Beach does not have any dune vegetation while the surveyed area in Boca Raton does. However, the results obtained with the chi-squared test indicated that the ratio of native and exotic species was not significantly different between the two areas. The p-value was 0.32, which is greater than the critical alpha value of 0.05 (Table 13). The frequency of native and exotic species surveyed in Pompano Beach and Boca Raton is given by the bar graph below

(Figure 21). When comparing the number of sites where Solenopsis invicta was detected, the two areas were not significantly different. The p-value was 0.48, which is greater than the critical alpha value of 0.05 (Table 14). The frequency of Solenopsis invicta in

Pompano Beach and Boca Raton is given by the bar graph below (Figure 22).

Table 13: Observed (O) and Expected (E) number of sites with native and exotic species

in Pompano Beach and Boca Raton (P>0.05)

Observed

Natives Exotics Total Surveyed Sites

Pompano Beach 6 6 12

Boca Raton 4 9 12

Grand Total 10 15 24

Expected

Natives Exotics Total Surveyed Sites

Pompano Beach 5 7.5 12

Boca Raton 5 7.5 12

Grand Total 10 15 24

p-value

0.32

Frequency of Native and Exotic Species per number of sites - Pompano Beach, FL vs. Boca Raton, FL

Exotics

Boca Raton Species

Natives Pompano Beach

0 2 4 6 8 10 Number of Sites

Figure 21: Graph of frequency of native and exotic species in Pompano Beach and Boca

Raton

Table 14: Observed (O) and Expected (E) number of sites with Solenopsis invicta in

Pompano Beach and Boca Raton survey areas (P>0.05)

Observed

S. invicta Total Surveyed

Sites

Pompano Beach 5 12

Boca Raton 3 12

Grand Total 8 24

Expected

S. invicta Total Surveyed

Sites

Pompano Beach 4 12

Boca Raton 4 12

Grand Total 8 24

p-value

0.48

Frequency of Solenopsis invicta in Pompano Beach and Boca Raton

Boca Raton

Pompano Beach Species Pompano Beach Boca Raton

0 2 4 6 Number of Sites

Figure 22: Graph of frequency of Solenopsis invicta in Pompano Beach and Boca Raton,

The survey conducted in Boca Raton was replicated four times to determine consistency in surveying methods. Although some variation is expected, the overall outcome was not expected to vary significantly among replicate surveys. The results obtained with the chi-squared test indicated that there is no significant difference in the ratio of native and exotic species among the four replicate surveys. The p-value was 0.70, which is greater than the critical alpha value of 0.05 (Table 15). The frequency of native and exotic species in Boca Raton for all four replicates is given by the bar graph below

(Figure 23). When comparing the number of sites where Solenopsis invicta was detected, the four replicates did not indicate significant differences. The p-value was 0.63, which is greater than the alpha critical value of 0.05 (Table 16). The frequency of Solenopsis invicta when comparing the four replicates is given by the bar graph below (Figure 24).

Table 15: Observed (O) and Expected (E) number of sites with Solenopsis invicta in

Pompano Beach and Boca Raton survey areas (P>0.05)

Observed

Natives Exotics Total Surveyed Sites

BR1 4 9 12

BR2 4 8 12

BR3 5 9 12

BR4 7 10 12

Grand 20 36 48

Total

Expected

Natives Exotics Total Surveyed Sites

BR1 5 9 12

BR2 5 9 12

BR3 5 9 12

BR4 5 9 12

Grand 20 36 48

Total

p-value

0.70

Frequency of Native and Exotic Species per number of sites - Pompano Beach, FL vs. Boca Raton, FL

BR4

BR3

Exotics

Replicates BR2 Natives

BR1

0 2 4 6 8 10 12 Number of Sites

Figure 23: Graph of frequency of native and exotic species in Boca Raton

Table 16: Observed (O) and Expected (E) number of sites with Solenopsis invicta in the

Boca Raton survey area – four replicates (P>0.05)

Observed

S. invicta Total Surveyed Sites

BR1 3 12

BR2 1 12

BR3 4 12

BR4 3 12

Grand Total 11 48

Expected

S. invicta Total Surveyed Sites

BR1 2.75 12

BR2 2.75 12

BR3 2.75 12

BR4 2.75 12

Grand Total 11 48

p-value

0.63

Frequency of Solenopsis invicta in Boca Raton

BR4

BR3 BR1 BR2

Replicates BR2 BR3 BR4 BR1

0 1 2 3 4 5 Number of Sites

Figure 24: Graph of frequency of Solenopsis invicta in Boca Raton - four replicates

6 FINAL REMARKS

The red imported fire ants impact a wide range of organisms, from native ant communities to endangered species such as the sea turtles nesting in South Florida and the U.S Virgin Islands. Mitigation efforts are required in order to control the spread of the species in areas where it is present, and avoid introduction to areas where the species is currently absent, like Jack Bay, St. Croix, USVI.

Efforts to control the RIFA’s impact on sea turtle hatchlings are on-going. The

Sea Turtle Research Team at Gumbo Limbo Nature Center periodically treats for fire ants if they are detected near sea turtle nests. The U.S Fish and Wildlife office in St. Croix also monitors for fire ant attacks on hatchlings.

However, a more concerted effort is needed in accurately identifying fire ants and determining whether or not the species is dominant in the area. The study of the distribution of an invasive species such as the RIFA can serve as a baseline to determine critical areas that need to be tackled in terms of control as well as areas that need to be monitored to keep these invasive ants from spreading. Knowing where the fire ants currently are can provide insight for future range expansions.

The methodology used in these surveys is the one most commonly used when surveying for dominant species of ants. Although variations on species detected occur, the likelihood of species found in one survey of an area not to be there in subsequent surveys is small. Moreover, there is a need to conduct baseline ant distribution surveys in other areas where sea turtles nest because – with globalization and climate change – it is likely that Solenopsis invicta will be introduced to other sea turtle nesting beaches.

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