By Mirabai R. Mccarthy the Climate and Structural Complexity of Tropical

Home , Adiantum tenerum, Phlebodium, Phlebodium aureum, Platycerium bifurcatum, Pleopeltis

ABSTRACT

BRYOPHYTE INFLUENCE ON TERRESTRIAL AND EPIPHYTIC FERN GAMETOPHYTES

by Mirabai R. McCarthy

The climate and structural complexity of tropical forests enable vertical habitat differentiation between epiphytic and terrestrial communities. Because fern spores are copiously released one might expect to find species growing in both habitats, yet taxa are typically confined to one particular habitat type. The goal of this study was to better understand how development of epiphytic and terrestrial fern species is restricted by habitat, by observing gametophyte growth on various substrates. Experiments revealed that terrestrial gametophytes were malformed, significantly reduced in size and did not reach sexual maturity or failed to germinate when sown directly on top of, or in close proximity to epiphytic bryophyte substrates. In contrast, epiphytic species developed uniformly regardless of distance to bryophyte substrates. These observations indicate that epiphytic bryophytes have a negative impact on terrestrial gametophyte development, perhaps through allelopathy, and this interaction appears to represent a limiting factor for the establishment of terrestrial species in epiphytic communities.

BRYOPHYTE INFLUENCE ON TERRESTRIAL AND EPIPHYTIC FERN GAMETOPHYTES

A Thesis

Submitted to the

Faculty of Miami University

in partial fulfillment of

the requirements for the degree of

Master of Science

Department of Botany

Mirabai Rachel McCarthy

Miami University, Oxford, Ohio

2007

Advisor______R. James Hickey

Reader______John Z. Kiss

Reader______Nicolas P. Money

TABLE OF CONTENTS

Introduction…………………………..……………………………..………………….….1

Materials and methods for substrate cultures………...... ….……….………...... 6

Materials and methods for substrate/agar cultures…..………..………………….…...... 11

Results from substrate cultures..…...……..……………………………...... …….…12

Results from substrate/agar cultures…..………………..………………………...……...24

Discussion………………..……………………………………………………...…….…33

Literature cited…………………..…………………………..…………………...……....38

LIST OF TABLES

Table 1. Collection and voucher data for plant material used in this study……………….8

Table 2. Mean developmental gametophyte stages and P-values……………………….20

iii LIST OF FIGURES

Figure 1. Map of Andros Island………………..………………………………...... 7

Figure 2. Stages of gametophyte development…..……………………………………...10

Figure 3. Phlebodium aureum and Neurodium lanceolatum development.…………….14

Figure 4. Platycerium bifurcatum and Thelypteris augescens development...... 16

Figure 5. Adiantum tenerum and Pteris bahamensis development……………...... 18

Figure 6. Development on hepatic and epiphytic soil substrates………….....………….19

Figure 7. Mean gametophyte stages on various substrates …...... 22

Figure 8. Epiphytic vs. terrestrial mean gametophyte stages on various

substrates……………………………………………………………………...23

Figure 9. Proportion of gametophyte stages across distances on hepatic/agar

cultures..…………………………………..….………………..……………...25

Figure 10. Adiantum tenerum and Neurodium lanceolatum development on

hepatic/agar cultures...... …...... ………………..………….………………….26

Figure 11. Proportion of gametophyte stages across distances on moss/agar

cultures……..………………………………………………...……………....27

Figure 12. Phlebodium aureum and Adiantum tenerum development on

moss/agar cultures………………………………………….………………...28

Figure 13. Mean gametophyte sizes on agar………………..…………………………...29

Figure 14. Mean gametophyte sizes on moss……………………..……………………..29

Figure 15. Proportion of gametophyte stages across distances on lichen/agar

cultures…………………..………………………………………………...…31

iv Figure 16. Asplenium nidus and Pteris bahamensis development on lichen/agar

cultures…………………..…..………………………….…………………....32

v ACKNOWLEDGEMENTS

I would like to thank…

Barb Wilson and Vickie Sandlin for absolutely everything you do. You are the glue that holds us all together!

Barbara Thiers, at the New York Botanical Garden and Bruce Allen, at the Missouri Botanical Garden, for their time and help with identification of moss and liverwort species.

The Department of Botany at Miami University, and the Ohio Academic Challenge grant program for providing financial support for my research.

Dr. Nic Money and Dr. John Kiss for their time and excellent feedback about my research.

All of my wonderful friends in the botany department, especially my lab mates, Susan Sprunt and Melanie Link-Perez, for their encouragement and support. You have all made my time here so much more enjoyable by sharing your passion for botany, some great laughs and unforgettable experiences! Thanks for bringing the feeling of a community into the department.

My mountain biking buddies in Heuston Woods. I seriously don’t think I would have survived life in Ohio without your company on the trails. Biking provides the perfect outlet for stress, and the opportunity to clear my thoughts. Your friendships have helped motivate me to get out there and ride. Thanks for all the great times, and the scars to remind me!

Dave Conant for sparking my interest in botany, and particularly ferns! His encouragement and guidance have greatly influenced my decision to pursue graduate school, and ultimately my direction in life. Thank you for being an excellent mentor, friend and role model.

My family; especially my mom for providing love and support every step of the way. Thanks for keeping me grounded and focused on the things that truly matter in life. I love you so much!

I would especially like to thank Jim Hickey for taking the time to provide thoughtful advice about my research, helping strengthen my knowledge about ferns, and further develop my passion for botany. Despite all of the jokes about your “nurturing” capabilities, you really have been a tremendous support and an excellent advisor! I sincerely appreciate the freedom you’ve granted me in the process of creating and exploring my research. That freedom has allowed me to develop a sense of independence and confidence about my abilities as a researcher and teacher. I can’t thank you enough for all that you do!

INTRODUCTION

The unique climate and structural complexity of tropical forests enable vertical habitat differentiation between terrestrial and epiphytic plant communities. Understanding the relationship between habitat type and species type, and when or how habitat selection occurs have been subjects of great interest and debate (Ranal, 1999; Hietz, et al., 2002; Cardelus et al., 2006; Watkins et al., 2007). Variations in plant morphology, species ranges, and community structure are the products of complex interactions among plants, animals, microbes and their surroundings (Ponge, 2002); however, the dynamics of these interactions are poorly understood. Tropical ecosystems contribute significantly to biodiversity worldwide. Epiphytes encompass about 10% of all vascular plants (Nieder et al., 2001; Callaway et al., 2002), of which pteridophytes are the second largest group (Hooper and Haufler, 1997). Epiphytes are either holo-epiphytic (where they grow on a host tree for their entire life cycle) or hemi-epiphytic (where a portion of their life cycle occurs on the ground; Nieder et al., 2001). Hemi-epiphytes are the least common form of epiphytism and account for less than 1% of all vascular plants (Nieder et al., 2001). In this study, the term “epiphyte” is used in a restrictive sense to include only holo-epiphytes. Species typically inhabit strictly epiphytic or terrestrial communities, but the ecological separation between habitats is not always clear. Occasionally species are found growing in opposing habitats, which implies that the mechanisms controlling habitat selection may not always be as stringent, or perhaps do not limit both gametophyte and sporophyte generations. Sillett (1999), and Mehltreter et al. (2005) discovered terrestrial fern species growing at the base of trees and in tree crotches where deep layers of humus have formed. These ferns were found exclusively in their juvenile form, and it was suggested that they remained fixed in the juvenile form as a response to growth in the wrong habitat (Mehltreter et al., 2005). Epiphytic ferns have been grown successfully from spore to sporophyte on manufactured terrestrial humus in a laboratory (Page,

2002), but they are rarely found growing on terrestrial soil in nature. These observations (Page, 2002) indicate that mechanisms preventing the establishment of epiphytic ferns in terrestrial communities are not likely the result of nutrient deficiencies in soil substrates, but rather are the result of other mechanisms in natural terrestrial communities, such as biochemical influences, attack by fungi or microherbivores, or space limitations. Epiphytic and terrestrial ferns rarely become established in alternate habitats, and the mechanisms that control their distribution are largely unknown. Epiphytism has evolved from terrestrial ancestors numerous times and in numerous families (Benzing, 1987), possibly as early as the Eocene era in ferns (Poole and Page, 2000). Although epiphytic ferns lack a common phylogenetic history, they do share some common traits. Most epiphytic fern species are members of the Polypodiaceae, Elaphoglossaceae, Vittariaceae, Grammitidaceae, and Hymenophyllaceae, five of the most derived fern families. Many sporophytes in these families have a climbing habit with surface clinging ability, reduced root systems, and many have rhizomes that are fine to filiform with long internodes (Poole and Page, 2000; Dubuisson et al., 2003). In addition, the gametophytes of epiphytic species are more adaptable and have a more plastic thallus (Stokey, 1951). Some adaptations common among epiphytic gametophytes include more extensive branching, more indeterminate growth, and a longer-lived life span compared to terrestrial gametophytes (Stokey, 1951; Atkinson, 1973; Farrar, 2003; Watkins et al., 2007). Additionally, many epiphytic gametophytes are able to produce clones or dispersible gemmae and can withstand extended periods of desiccation (Ong and Ng, 1998; Poole and Page, 2000; Dassler and Farrar, 2001; Page, 2002; Chiou et al., 2002; Farrar, 2003; Watkins et al., 2005; Watkins et al., 2007). Selective pressures vary across epiphytic and terrestrial habitats (Chiou and Farrar, 1997). Potential benefits associated with terrestrial habitats include an abundance of space, water and nutrients, though this may vary among tropical forests (Vitousek and Sanford, 1986). Benefits associated with epiphytic habitats are reduced predation (Hamilton and Pryor, 1994), less likelihood of disturbance

(by elevating fronds off the forest floor) (Benzing and Atwood, 1984; Hooper and Haufler, 1997), and decreased competition (Hamilton and Pryor, 1994). Another possible benefit of epiphytic habitats, and one of particular interest in this study, is the apparent relationship between epiphytic bryophytes and ferns. Many bryophyte species inhibit angiosperm growth via allelopathy (Asakawa, 1994; Zamfir, 2000; Sedia and Ehrenfeld, 2003), and some bryophytes possess antibiotic (polyphenolic compounds, cyclopental fatty acids and their precursors, Basile et al., 1998) and antifungal properties (Banerjee and Sen, 1979). Some reports suggest that these compounds contribute to species survival in extreme habitats, and provide protection from pathogens and herbivores (Onyilagha and Grotewold, 2004). Frequently bryophytes and fern gametophytes are found growing in association with one another (Pessin, 1925; Dassler and Farrar, 2001; Zotz and Hietz, 2001; Hsu et al., 2002; Dubuisson et al., 2003; Ellyson and Sillett, 2003). According to Dassler and Farrar (2001), epiphytic gametophytes are able to compete with bryophytes because they have a similar growth form. Mosses, liverworts, and lichens are commonly found in epiphytic habitats (Dassler and Farrar, 2001; Ellyson and Sillett, 2003), and according to Sedia and Ehrenfeld (2003), they play an integral role in plant community establishment. Potential limitations associated with terrestrial habitats include disturbance by ground dwelling mammals, increased predation (Hamilton and Pryor, 1994), and increased competition for space (Nieder et al., 2001). Previous research has identified several factors as restrictions for growth in epiphytic habitats. The first such limitation is nutrient disequilibrium. For example, some studies (Zotz and Hietz, 2001; Hietz et al., 2002; Hsu et al., 2002) suggest that nitrogen availability is limited in epiphytes. Some fern species have developed ways to overcome this limitation by obtaining nutrients from the atmosphere, or from decaying leaf litter (Hietz et al., 2002), whereas others such as Vittaria, Mecodium, Elaphoglossum, and Huperzia form symbiotic relationships with mycorrhizal fungi, which are able to assist with nutrient uptake (Gemma and Koske, 1995). A second perceived limitation is longevity of habitat. Epiphytic habitats are believed to be ephemeral because of tree falls, wind breaks, sloughing of bark and branches, and

disturbance by tree dwelling animals (Hooper and Haufler, 1997; Ong and Ng, 1998; Lyons et al., 2000; Zotz and Hietz, 2001). The lifespan of an epiphyte is constrained by the survival of its host. A third limitation of the epiphytic habitat is the constraint that it imposes upon reproductive behavior, such as proximity of conspecific gametophytes. According to Chiou and Farrar (1997), Chiou et al. (1998, 2002), and Farrar, (2003), epiphytic gametophytes in the Polypodiaceae and Elaphoglassaceae are capable of indeterminate growth, allowing the gametophyte to grow across rough terrain and through masses of bryophytes to other gametophytes. Limiting the distance between potentially antheridiate and archegoniate gametophytes thus increases the chances of fertilization and outcrossing. This is important because in most species there is a temporal separation between the production of gametangia types – dichogamy (Hooper and Haufler, 1997). A final limitation in epiphytic habitats is water availability (Benzing, 1987; Ong and Ng, 1998; Greer and McCarthy, 1999; Kentner and Mesler, 2000; Zotz and Hietz, 2001; Page, 2002). According to Benzing (1987), water availability is the most influential abiotic factor affecting survival in epiphytic habitats. Most epiphytic substrates lack the ability to retain water, thereby creating an inconsistent moisture supply and frequent periods of desiccation. In experiments conducted by Watkins et al. (2005), gametophytes of epilithic and epiphytic ferns were found to have excellent abilities to recover from desiccation. Epiphytic gametophytes were able to tolerate 16 - 25 months of desiccation, whereas terrestrial gametophytes were only able to tolerate 4 - 11 months (Watkins et al., 2005). Survival after prolonged periods of desiccation suggests a strong correlation between epiphytic gametophyte recovery and their ability to grow in arid conditions. It is unclear specifically how, or to what degree, niches vary between gametophyte and sporophyte generations. However, studies by Tryon and Vitale (1977) indicate that gametophytes appear to occupy a more selective niche, particularly terrestrial gametophytes, which are often found growing in moist microhabitats within a parent population, or in recently disturbed areas (Benzing and Atwood, 1984). Potentially, habitat selection could occur at any stage of the

life cycle because environmental pressures are constantly being applied, but according to Page (2002) the greatest pressure is presumably exerted on the gametophyte generation. Page (2002) also describes the independent gametophyte as a handicap because it will not tolerate widely fluctuating conditions, its rate of growth is slow, and it is critically dependent on water. However, recent studies suggest that gametophytes are more prevalent than sporophytes and reportedly have the ability to tolerate a greater amount of stress (Watkins et al., 2007). Due in part, to this apparent divergence in understanding about which stage in pteridophyte development is most sensitive to selective pressures, this study chose to focus on the gametophyte generation. Ferns produce an abundance of spores, which are dispersed by wind into both epiphytic and terrestrial communities. The majority of spores are deposited in the vicinity of parent plants (Ramirez-Trejo et al., 2004), but transport of spores into opposing habitats can occur by wind (Dassler and Farrar, 2001). Spores are incredibly resistant to extremes in temperature, moisture, altitude, and irradiation (Page, 2002), and reportedly many have the ability to remain viable for decades (Ramirez-Trejo et al., 2004). Ranal (2004), analyzed the bark of several angiosperm tree species and found an accumulation of viable spores from fourteen terrestrial and one epiphytic species. Despite the abundance of terrestrial spores on the bark, there was no indication of gametophyte growth anywhere on the trees. Ramirez-Trejo et al. (2004) discovered several different species of viable, epiphytic spores in a soil spore bank, which upon placement into controlled growth conditions germinated on the soil. The ability of spores to germinate in opposing habitats is clearly impeded by unknown environmental constraints (Ramirez-Trejo et al., 2004; Ranal, 2004). Previous studies (Stokey, 1951; Nayer and Kaur, 1971; Benzing, 1987; Gemma and Koske, 1995; Ong and Ng, 1998; Poole and Page, 2000; Nieder et al., 2001; Hietz et al., 2002; Watkins et al., 2005) provide some insight into how tropical ferns have adapted to their unique environments, and suggest that a restriction exists. However, understanding when or how the selection against an epiphytic or terrestrial community occurs is beyond the scope of these studies. Research has

not yet explained how ferns of epiphytic and terrestrial communities remain exclusive to their habitat when spores are present and viable across habitats. In an effort to better understand habitat limitations, this study attempts to identify whether development of epiphytic and terrestrial gametophytes is negatively impacted when both species types are grown on substrates from opposing epiphytic and terrestrial habitats.

MATERIALS AND METHODS

Three terrestrial fern species Adiantum tenerum Sw., Thelypteris augescens (Link) Munz and I. M. Johnst., Pteris bahamensis (Ag.) Hieron., and three epiphytic species Phlebodium aureum L., Neurodium lanceolatum (L.) Fee., Campyloneuron phyllitidis (L.) Presl. were collected from various sites on the island of North Andros in the Bahamas (Fig. 1) in May 2006, and vouchers are housed at Miami University (Table 1). Fertile portions of the fronds were placed in newspaper and allowed to dry. Dehisced spores were transferred to individual sealed envelopes and stored for later use. Upon removal from envelopes, spores were sifted through kim wipes ® to separate spores from sporangial debris. Several epiphytic substrates, including two hepatic species, Cheilolejeunea rigidula (Nees. and Mont.) R.M. Schust., and Lejeunea flava (Sw.) Nees., one moss species, Calymperes palisotii Schwaegr., an unknown lichen species, and epiphytic soil were also collected. We were unable to obtain a collecting permit for terrestrial soil in the Bahamas. Instead, similar limestone rich soil was obtained from southern Florida to be utilized as a terrestrial substrate. Additional epiphytic ferns, Pyrrosia piloselloides (L.) M. Price., Asplenium nidus L., and Platycerium bifurcatum (Car.) C. Christens., were obtained from the Miami University Greenhouse, and spores were collected in the same manner as listed above.

SUBSTRATE CULTURES - Spores of three epiphytic fern species, Phlebodium aureum, Neurodium lanceolatum, Platycerium bifurcatum, and three terrestrial fern species, Adiantum tenerum, Thelypteris augescens, Pteris bahamensis were

FIG. 1. Map of Andros Island in the Bahamas. All fern, lichen, and bryophyte specimens were collected in N. Andros.

TABLE 1. Collection and voucher data for plant material used in this study.

Species Collection Data

Adiantum Maidenhair coppice. Collected in disturbed habitat, along tenerum roadside in sandy limestone soil, partial shade. Near, N. 24°, 47.913’, W. 77°, 53.420’. McCarthy 13 (MU)

Thelypteris BARC roadside. Growing in disturbed habitat, high light, in augescens limestone soil. N. 24°, 58.113’, W. 78°, 51.382’. McCarthy 06 (MU)

Pteris Near Goby Lake. Collected at base of turned up tree in dry, bahamensis sandy, limestone soil in high light. N. 24°, 49.699’, W. 77°, 55.379’. McCarthy 17 (MU)

Phlebodium Jungle Pond. Epiphytic on Rhizophora mangle about 6 ft above aureum ground in shade. McCarthy 08 (MU)

Neurodium Maidenhair Coppice. Epiphytic on Lysaloma sabicu, about 3 ft lanceolatum above ground in partial shade. N 24°, 47.926’, W. 77°, 53.818’ McCarthy 09 (MU)

Campyloneuron Maidenhair Coppice. Epiphytic on Cocoloba diversifolia, about phyllitidis 1 foot above ground in epiphytic soil and partial shade. N 24°, 47.926, W. 77°, 53.818 McCarthy 03 (MU)

Pyrrosia Miami University, Boyd Greenhouse. piloselloides McCarthy 18 (MU)

Asplenium nidus Miami University, Boyd Greenhouse. McCarthy 20 (MU)

Platycerium Miami University, Boyd Greenhouse. bifurcatum McCarthy 19 (MU)

Lejeunea flava Jungle Pond. Epiphytic on Rhizophora sp. about 5 ft above ground, in association with Neurodium lanceolatum McCarthy 21 (MU)

Calymperes Maidenhair Coppice, epiphytic on Conocarpus erectus, in palisotii association with Polypodium polypodioides. McCarthy 22 (MU)

Flavoparmelia Maidenhair Coppice, epiphytic on Lysiloma sabicu, in carperata (?) association with Neurodium lanceolatum. McCarthy 23 (MU)

soaked in distilled water for 20 minutes, surface sterilized in a 5% commercial bleach solution (cfern.bio.utk.edu) and suspended in 5 ml of distilled water. Epiphytic substrates and terrestrial soil were autoclaved, then separately distributed into 30 mm Petri dishes. A Pasteur pipette was utilized to distribute three droplets of spores onto epiphytic soil, terrestrial soil, and hepatic substrates. As a control, spores were sown on agar (1.2 % w/v), fortified with Murashige and Skoog solution (www.jrhbio.com; Fernandez and Revilla, 2003). Six replicate Petri dishes per species, for each of the six media types, were sown under a laminar flow hood and sealed with Parafilm®. Petri dishes were put into Tupperware® containers (16” x 22”) and placed into a growth chamber at 20°C, under fluorescent light with a 12 hr photoperiod. Weekly observations were made using an Olympus SZX-12 dissecting microscope, and digital images were obtained using an Olympus 4600. Individual gametophyte development was scored by category as belonging to one of six stages (Fig. 2) and tracked over a period of 55 days. Populations of fifteen gametophytes were randomly selected from each of the Petri dishes, and mean gametophyte stage was determined for each species on each substrate after 55 days. Gametophyte development was compared to expected developmental progress as identified during preliminary observations and reports by Stokey and Atkinson (1954), Kato (1970), Atkinson (1971, 1973), Huckaby and Raghaven (1981), Camloh (1993), Chiou and Farrar (1997), Chiou et al. (2002), Ko (2003), Perez-Garcia and Mendoza-Ruiz (2004). Mean gametophyte stage was compared between species and habitat type on agar, epiphytic soil, terrestrial soil, and hepatic substrates. ANOVA was used to determine whether there were differences in gametophyte development, as assessed by mean stage (0-5), on the various substrates (agar, epiphytic soil, hepatic, terrestrial soil). A post hoc analysis, using Tukey pairwise comparisons, was later used to specifically identify the difference in means. Significant values were based upon a corrected Bonferroni p-value of 0.0008. Substrate effects on species type (epiphytic/terrestrial) were determined using a pairwise comparison with ANOVA. These statistical analyses were performed using SAS (SAS Institute, Inc. 2001).

FIG. 2. Stages of gametophyte development scored in current study. 0. Spore prior to germination. 1. Filamentous gametophyte with rhizoid. 2. Two dimensional growth. 3. Development of a notch meristem. 4. Development of sex organs (antheridia and/or archegonia). 5. Development of sporophyte.

EPIPHYTIC SUBSTRATE & AGAR CULTURES - Spores of three epiphytic fern species, Phlebodium aureum, Pyrrosia piloselloides, Asplenium nidus, and three terrestrial fern species Adiantum tenerum, Thelypteris augescens, and Pteris bahamensis were soaked in distilled water for 20 minutes and surface sterilized as described above. Epiphytic substrates were autoclaved, and then, under a laminar flow hood, each was separately combined with agar (1.2 % w/v, fortified with Murashige and Skoog solution (Fernandez and Revilla, 2003)) in Petri dishes, such that a piece of lichen, hepatic, or moss (each substrate approximately equal in size) was distributed and centrally embedded into 9 cm agar-filled dishes. Three replicates per species, per treatment were prepared. Spores were treated as described above and droplets of spores were sown on top of, and in concentric rings outward from the piece of substrate at 1 cm distances. The Petri dishes were sealed with parafilm and placed into Tupperware® containers (16” x 22”) in a growth chamber under the same conditions as described above. Gametophyte development was monitored over a span of 60 days by making weekly observations using an Olympus SZX-12, dissecting scope. Gametophyte development was categorized according to the stages defined in Fig. 2, for each species and substrate combination. Fifteen gametophytes were sampled and scored for developmental stage at each of six populations, at distances of 0, 1, 2, 3, and 4 cm from an epiphytic substrate source. Mean developmental stage was calculated at each of these distances. Gametophyte size was measured on moss/agar cultures using ImagePro software. Populations of 10 gametophytes from three epiphytic and three terrestrial species were measured directly on top of the moss, and at a distance 3 - 4 cm away from the moss substrate, on agar media. The gametophyte measurements were analyzed using ANOVA procedures.

RESULTS

SUBSTRATE CULTURES – Observations were recorded from a total of 2,160 gametophytes on four substrates (agar, epiphytic soil, hepatic, terrestrial soil). Gametophytes developed to stages 3 and 4 on agar and epiphytic soil in all species, except terrestrial Thelypteris augescens, which developed only to stage 2 on agar (Figs. 3 - 5). The terrestrial soil substrate negatively effected development of all species (Figs. 3 - 5). Gametophytes of terrestrial Adiantum tenerum, Pteris bahamensis, and Thelypteris augescens developed cordiform thalli (stage 3) on terrestrial soil, then died after about a month (Figs. 4 & 5). Epiphytic, Neurodium lanceolatum and Platycerium bifurcatum failed to germinate on terrestrial soil (Figs. 3 & 4), and epiphytic Phlebodium aureum developed to stage 1, then died after about a month (Fig. 3). Gametophyte mortality on terrestrial was so high that development on this substrate could not be determined with any degree of confidence, and was therefore dropped from further analysis. The hepatic substrate negatively effected development of the terrestrial species, but did not negatively impact epiphytic development (Figs. 3 – 5 & 6a & b). Terrestrial Adiantum tenerum, and Pteris bahamensis failed to germinate on hepatic substrate (Figs. 5 & 6b), and Thelypteris augescens only developed to stage 1. Thelypteris augescens gametophytes were also severely malformed. Development of epiphytic Neurodium lanceolatum, Phlebodium aureum and Platycerium bifurcatum on hepatic substrate was normal and relatively equal to development on other substrates (excluding terrestrial soil) (Figs. 3, 4 & 6a). During the growth period of 55 days Phlebodium aureum and Neurodium lanceolatum gametophytes developed to stage 3 on agar, epiphytic soil and hepatic substrates. Development of epiphytic Platycerium bifurcatum was exceptionally low on all substrates (Table 2). Previous studies indicate that Platycerium bifurcatum is expected to germinate on agar in 5-7 days and develop antheridia in 20-24 days (Fig. 4). Due to the exceptionally low germination and slow development of Platycerium bifurcatum across all substrates, it was dropped from further analysis.

Species Fig. 3. Gametophyte development, stages 0-5, on agar, epiphytic soil, hepatic, and terrestrial soil in Phlebodium aureum and Neurodium lanceolatum. Expected 1 2 3 times are those reported by Ko (2003), Chiou and Farrar (1997), and Chiou, Farrar, and Ranker (2002) on agar (a skull and cross bones symbol represents gametophyte mortality). Expected times for Neurodium lanceolatum were not found in the literature.

Phlebodium aureum

Agar

Epi Soil

Hepatic

Terr Soil

Expected 1,2,3 Times

Days from 0 5 10 15 20 25 30 35 40 45 50 55 Germination

Neurodium lanceolatum

Agar

Epi Soil

Hepatic

Terr Soil

Expected Times ?

Days from 0 5 10 15 20 25 30 35 40 45 50 55 Germination

Fig. 4. Gametophyte development, stages 0-5, on agar, epiphytic soil, hepatic, and terrestrial soil in Platycerium bifurcatum and Thelypteris augescens. Expected times are those reported by 1Stokey, and Atkinson (1954), 2Atkinson (1971), 3 Huckaby and Raghavan (1981), 4Camloh (1993), and 5Perez-Garcia and Mendoza-Ruiz (2004), on agar (a skull and cross bones symbol represents gametophyte mortality).

Platycerium bifurcatum

Agar

Epi Soil

Hepatic

Terr Soil

Expected 1,4 Times

Days from Germination 0 5 10 15 20 25 30 35 40 45 50 55

Thelypteris augescens

Agar

Epi Soil

Hepatic

Terr Soil

Expected 2,3,5 Times

Days from 0 5 10 15 20 25 30 35 40 45 50 55 Germination

Fig. 5. Gametophyte development, stages 0-5, on agar, epiphytic soil, hepatic, and terrestrial soil in Adiantum tenerum and Pteris bahamensis. Expected times are those reported by 1Kato (1970), and 2Atkinson (1973) on agar, and those discovered during preliminary growth experiments on agar (a skull and cross bones symbol represents gametophyte mortality).

Adiantum tenerum

Agar

Epi Soil

Hepatic

Terr Soil

Expected Times

Days from 0 5 10 15 20 25 30 35 40 45 50 55 Germination

Pteris bahamensis

Agar

Epi Soil

Hepatic

Terr Soil

Expected 1,2 Times

Days from 0 5 10 15 20 25 30 35 40 45 50 55 Germination

Hepatic Substrate Epiphytic Soil Substrate

FIG. 6. Epiphytic Phlebodium aureum (A), and terrestrial Pteris bahamensis (B) gametophytes on hepatic substrate (Lejeuna flava), and epiphytic soil substrate after 55 days. Scale bars are approximately 0.5 cm.

TABLE 2. Development of terrestrial and epiphytic gametophytes after 55 days on various substrates. The values given represent mean developmental stage (0-5), standard deviation, and range. Mean differences were identified using ANOVA (Significant P-values are based on a corrected Bonferroni of 0.0008). There was a significant difference in mean stages (Pr < 0.0001) on the various substrates in all species except Platycerium bifurcatum (Pr < 0.0386), and Neurodium lanceolatum (Pr < 0.0013).

Agar Epiphytic Hepatic Terrestrial Pr. Value Soil Soil Terrestrial Species

Thelypteris 0.74 +/- 2.61 +/- 0.03 +/- 0 +/- 0 F3,20 = augescens 0.80 0.665 0.180 (0-0) 112.2, (0-2) (1-4) (0-1) P = .0001

Pteris 1.67 +/- 2.26 +/- 0 +/- 0 0.13 +/- 0.5 F3,20 = bahamensis 0.99 0.73 (0-0) (0-2) 42.02, (1-4) (1-4) P = .0001

Adiantum 2.05 +/- 1.48 +/- 0 +/- 0 0 +/- 0 F3,20 = tenerum 0.91 0.91 (0-0) (0-0) 25.22, (0-4) (0-3) P = .0001

Epiphytic Species

Neurodium 1.05 +/- 1.48 +/- 1.16 +/- 0 +/- 0 F3,20 = 7.71, lanceolatum 0.46 0.75 0.91 (0-0) P = .0013 (0-2) (0-3) (0-3)

Phlebodium 2.57 +/- 1.82 +/- 1.10 +/- 0 +/- 0 F3,20 = aureum 0.77 0.86 0.83 (0-0) 22.33, P = (0-3) (1-3) (0-3) .0001

Platycerium 0.16 +/- 0.07 +/- 0 +/- 0 0 +/- 0 F3,20 = 20, bifurcatum 0.47 0.37 (0-0) (0-0) P = .0386 (0-2) (0-2)

Ninety gametophytes per species, per substrate were scored by stage (Fig. 2), and mean stages were determined after a 55-day growth period (Table 2 & Fig. 7). Results from ANOVA indicate that there was a significant difference (Pr < 0.0001) in development on agar, epiphytic soil, hepatic, and terrestrial soil substrates in all species except epiphytic Neurodium lanceolatum (Pr < 0.0013), based upon a Bonferroni corrected P-value of 0.0008 (Table 2). A post hoc analysis using Tukey pairwise comparisons indicated that mean terrestrial development of Adiantum tenerum and Pteris bahamensis gametophytes was significantly different on hepatic, than on agar or epiphytic soil substrates (Fig. 7). This difference was not significant in epiphytic Phlebodium aureum, or Neurodium lanceolatum (Fig. 7). Specifically, all terrestrial species, Adiantum tenerum, Pteris bahamensis, and Thelypteris augescens either failed to germinate on hepatic substrate, or were severely malformed. Thelypteris augescens (Figs. 4

& 7c) gametophytes developed to stage 1 on the hepatic substrate and remained locked at this stage throughout the duration of the study; the spores germinated, but never attained a two-dimensional form. Instead, they developed into globular masses of chlorophyllous cells. Germination of epiphytic Neurodium lanceolatum and Phlebodium aureum on hepatic substrate was normal, and showed germination rates comparable to that on other substrates (Fig. 7). Mean developmental data were pooled to form two categories; one for terrestrial species, the other for epiphytic species. A Bonferroni – adjusted pairwise comparison was used to check for differences in mean development between the pooled data by substrate. Based on an adjusted P-value of 0.0125, the analysis determined that significant differences (P < 0.0001) in development occurred on the hepatic substrate (Fig. 8).

ab a a ab b

c c c

A D

a a

b b

B E

c c

FIG. 7. Mean gametophyte development of terrestrial (A - C) and epiphytic (D - E) species on agar, epiphytic soil, hepatic, and terrestrial soil substrates. Letters above each box indicate significant differences between substrates as determined by Tukey’s pairwise comparisons using a Bonferroni corrected P-value of 0.0008. The difference in means of Neurodium was not significant.

Fig. 8. Mean gametophyte development of terrestrial and epiphytic (excluding Platycerium) species on agar, epiphytic soil (epi soil), hepatic, and terrestrial soil (terr soil) substrates after 55 days. Significant differences were found between epiphytic and terrestrial means on the hepatic substrate (Pr < 0.0001). Based upon a corrected Bonferroni P-value of 0.0125, the difference between epiphytic and terrestrial means on agar (Pr > 0.087), epiphytic soil (Pr > 0.0158, and terrestrial soil (Pr > 0.811) substrates was not significant.

EPIPHYTIC SUBSTRATE/AGAR CULTURES - Gametophytes of terrestrial Thelypteris augescens, Pteris bahamensis, and Adiantum tenerum stage were more likely to develop beyond stages 0-1 as the distance from hepatic substrate increased (Fig. 9a – c). Spores of these species (Thelypteris augescens, Pteris bahamensis, and Adiantum tenerum) located directly on top of, or in proximity to the hepatic point source, either failed to germinate (Fig. 10a), or began to germinate and developed into globular, irregularly shaped gametophytes (Fig. 10b). At increasing distances from the hepatic point source, two-dimensional gametophytes were found, but they still showed some malformation (Fig 10c). In contrast, epiphytic Neurodium lanceolatum and Phlebodium aurem gametophytes developed in a relatively uniform fashion regardless of distance from the hepatic point source (Figs. 9d - e & 10e); stage 3 cordiform thalli were found at all distances. On the moss/agar cultures terrestrial gametophyte development was positively correlated with distance from the moss (Figs. 11a - c & 12b). Terrestrial Thelypteris augescens, Pteris bahamensis and Adiantum tenerum populations had greater proportions of earlier stage (stage 2), pre-cordate gametophytes both on, and around the moss point source, and the proportion of later stage gametophytes (stages 3 & 4) increased with distance from the moss (Fig. 11a - c). In contrast, the epiphytic species Phlebodium aurem, Pyrrosia piloselloides, and Asplenium nidus showed relatively homogeneous gametophyte development across distances B. (Figs. 11d – f & 12a) on the moss/agar substrates. At nearly all distances, epiphytic gametophyte populations were composed of cordiform thalli bearing sex organs (stages 3 & 4), with some sporophyte (stage 5) development intermixed across all distances (Fig. 12). Differences in gametophyte size were observed in terrestrial species on moss/agar cultures. Measurements taken from gametophytes directly on agar substrate showed no significant difference (P> 0.05) in size among or between terrestrial and epiphytic species (Fig. 13). However, measurements taken from epiphytic and terrestrial gametophytes on moss were significantly different in size

24 Adiantum tenerum Neurodium lanceolatum 100%

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Thelypteris augescens Phlebodium aureum 100%

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0% B. 0 1 2 3 4 5 Distance Fom Hepatic (cm) E. Pteris bahamensis 100%

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0% 0 1 2 3 4 5

Distance From Hepatic (cm) C.

Stage 0 Stage 1 Stage 2 Stage 3 Stage 4 Stage 5

Fig. 9. Proportion of terrestrial (A - C) and epiphytic (D & E) gametophytes at various developmental stages (0-5), at distances of 0-5 cm from epiphytic hepatic point source.

B C

FIG. 10. Development of terrestrial, Adiantum tenerum (A – C), and epiphytic Neurodium lanceolatum (D) (scale bar approximately 50mm) on agar/hepatic cultures after 55 days. (A) A. tenerum spores (approximately 40 µm) sown directly on top of hepatic point source failed to germinate. (B) Spores sown a short distance from point source developed into malformed globular gametophytes. (C) At further distances from hepatic, gametophytes grew a bit larger, but were still malformed. (D) N. lanceolatum developed normally at all distances from hepatic.

Adiantum tenerum Phlebodium aureum 100% 90% 80% 70% 60% 50% 40% 30% 20% 10% 0% A. D. Thelypteris augescens Pyrrosia piloselloides 100%

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0% B. E. Asplenium nidus 100% Pteris bahamensis

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0% 0 1 2 3 4 0 1 2 3 4 Distance From Moss (cm) C. Distance From Moss (cm) F.

Stage 0 Stage 1 Stage 2 Stage 3 Stage 4 Stage 5

Fig. 11. Proportion of terrestrial (A-C) and epiphytic (D-F) gametophytes at various developmental stages (0-5), at distances of 0-5 cm from epiphytic moss point source.

FIG. 12. Epiphytic Phlebodium aureum (A) and terrestrial Adiantum tenerum (B) gametophytes on agar/moss (Calymperes palisotii) cultures after 55 days (scale bars are approximately 1 cm).

FIG. 13. Mean gametophyte size and SD of epiphytic (Lemmaphyllum, Asplenium, Phlebodium) and terrestrial (Adiantum, Thelypteris, Pteris) species on agar substrate.

FIG. 14. Mean gametophyte size and SD of epiphytic (Lemmaphyllum, Asplenium, Phlebodium) and terrestrial (Adiantum, Thelypteris, Pteris) species on moss substrate.

among species (F5,54 = 62.68, P < 0.0001) (Fig. 14). Terrestrial Thelypteris augescens, Pteris bahamensis and Adiantum tenerum gametophytes averaged between 2 & 10 times smaller than epiphytic Phlebodium aurem, Pyrrosia piloselloides, and Asplenium nidus gametophytes on moss substrates (Fig. 14). On lichen/agar cultures all species developed to cordiform (stage 3), and sexual (stage 4) gametophytes at all distances (Figs. 15 & 16). Small percentages of stage 0 & 1 gametophytes were present in all species, between 0 – 1 cm away from the lichen point source. Gametophyte stage was generally uniform across all distances in both epiphytic and terrestrial species. There was no perceptible correlation between species type (epiphytic/terrestrial) and distance from the lichen point source (Figs. 15 & 16).

30 Adiantum tenerum Phlebodium aureum 100%

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Stage 0 Stage 1 Stage 2 Stage 3 Stage 4 Stage 5

Fig. 15. Proportion of terrestrial (A-C) and epiphytic (D-F) gametophytes at various developmental stages (0-5), at distances of 0-5 cm from epiphytic lichen point source.

FIG. 15. Epiphytic Asplenium nidus (A), and terrestrial Pteris bahamnesis (B) gametophytes on lichen/agar cultures after 55 days (scale bars are approximately 1 cm).

DISCUSSION Benzing (1987) discussed potential reasons why some ferns are epiphytic and others are not. He concluded that epiphytes have a thick cuticle, strictly regulate water, have low transpiration and photosynthetic rates, and possess a tolerance to desiccation. Benzing (1987) suggested that more data on substrates and microclimates for both sporophyte and gametophyte generations was necessary to better understand the evolution of pteridophyte epiphytism. Paoletti et al. (1991), found that despite similar decomposition rates, epiphytic soils had higher concentrations of carbon and minerals than terrestrial soils. Watkins et al. (2005) compared germination rates of terrestrial and epiphytic ferns on epiphytic and terrestrial soils and discovered that, under laboratory conditions, germination was 60% higher on epiphytic soils. In the present study terrestrial gametophyte germination was greater on epiphytic soil than on any other substrate (Fig. 8). These observations, as well as those made by Watkins et al. (2005) show that, in nature, growth of terrestrial species on epiphytic soils may be restricted by factors other than nutrient availability (due to the fact that growth occurs normally on epiphytic soil in a laboratory). In natural habitats growth of terrestrial species on epiphytic soils may be limited by water accessibility, gametophyte morphology, epiphytic pathogens, and biochemical influences (Chiou and Farrar, 1997; Chiou et al., 1998, 2002; Onyilagha and Grotewold, 2004; Watkins et al., 2005). In the current study, development of gametophytes from all terrestrial species was extremely poor when grown directly on top of, or in close proximity to epiphytic bryophytes. This curious pattern was observed in terrestrial gametophytes that were grown on hepatic (Fig. 6b), hepatic/agar (Fig. 10a - c), and moss/agar (Fig. 12b), substrates. Epiphytic gametophytes of Phlebodium aureum, and Neurodium lanceolatum developed normally when grown directly on hepatic substrates (Fig. 6a), and epiphytic gametophytes of Phlebodium aureum, Asplenium nidus, and Pyrrosia pilloselloides developed normally when grown on hepatic/agar (Fig. 10) and moss/agar (Fig. 12a) substrates. All gametophytes developed normally on agar, epiphytic soil (Fig. 6), and lichen/agar (Fig. 16) substrates.

Epiphytic and terrestrial gametophyte development was essentially homogenous on all substrates, except those that were composed of epiphytic bryophytes; whereby development of terrestrial species was severely hindered in the presence of mosses and hepatics. These data indicate that substrate can influence gametophyte development. However, habitat-specific substrates (i.e., substrates collected strictly from epiphytic communities) do not appear to have a blanket negative effect on development in species from opposing habitats. The epiphytic lichen and epiphytic soil did not negatively affect terrestrial development. This suggests that bryophytes may represent the primary roadblock to establishment of terrestrial fern species in epiphytic habitats. The architecture of epiphytic surfaces consists of an interwoven assemblage of bark, lichens, and bryophytes. Lichens and bryophytes are particularly dominant occupants of epiphytic habitats (Roberts et al., 2005), and numerous studies have reported examples of bryophytes and ferns growing in association with each other (Pessin, 1925; Sillett, 1999; Dassler and Farrar, 2001; Hsu et al., 2002; Ellyson and Sillett, 2003; Dubuisson et al., 2003; Roberts et al., 2005). Studies by Sedia and Ehrenfeld (2003) found that moss and lichen play an essential role in community establishment, possibly because bryophytes are ectohydric and are capable of improving water retention (Zamfir, 2000). Competition among bryophytes and epiphytic ferns is presumably limited as evidenced by the frequent overlap in niche occupancy. In fact, it seems possible that epiphytic ferns may even benefit from cohabiting with bryophytes. For example, aromatic compounds and terpenoids, which are commonly found in liverworts (Asakawa, 1994) are reportedly biologically active against attack by bacteria, fungi, and parasitic microorganisms, and may induce dermatitis in humans, and inhibit plant growth via allelopathy (Banerjee and Sen, 1979; Asakawa, 1994). Lichens and mosses have also demonstrated antifungal, antibacterial (Banerjee and Sen, 1979), and allelopathic effects on vascular plant development (Zamfir, 2000; Sedia and Ehrenfeld, 2003).

Allelopathic interactions are the result of inhibitory plant chemicals that are found in stems, leaves, roots, flowers (including pollen, fruits, and seeds) and epicuticular wax (Bell and Klikoff, 1979). Allelochemicals, are derived from secondary metabolites, such as phenolic acids, coumarins, alkaloids, terpenoids, and flavonoids (Onyilagha and Grotewold, 2004). They are secreted from plants into soil, via leaching, decomposition, or root exudation, and can affect the early phases of plant development (An, 2005). Allelopathy has been targeted as an ecological factor determining plant dominance, succession, species diversity, and community structure (An, 2005). Reports suggest that flavonoids and phenols exhibit anitibacterial, anti-inflamatory, and antiviral properties, and contribute to blocking infection, herbivory defense, and reducing pathogens in host plants (Alonso-Amelot et al., 2004; Onyilagha and Grotewold, 2004). Flavonoids may also shield plants from the damaging effects of ultraviolet radiation, and play an important role in species survival in extreme conditions (Onyilagha and Grotewold, 2004). Recently, 112 weed species were listed as having allelopathic properties (Foy and Inderjit, 2001). These discoveries suggest that plants armed with allelopathic compounds make better competitors and might have the ability to develop invasive tendencies as a result of their armament. Most research on allelopathy has centered on angiosperms, primarily because of their importance in crop production and weed control. However, allelopathy has been documented in bryophytes, lichens, and ferns (Asakawa, 1994; Zamfir, 2000; Sedia and Ehrenfeld, 2003; Munther and Fairbrothers, 2006). Phytochemicals are pervasive in ferns, (Page, 2002). Compounds such as cinnamomic acid, flavonoids, and thelypterins are found in both gametophyte and sporophyte generations (Lloyd, 1974; Wagner, 1974; Page, 2002; Munther and Fairbrothers, 2006). It has been hypothesized that sporophytes inhibit gametophyte growth because the gametophyte generation is rarely found growing directly under sporophyte populations in the field or in culture plants (Munther and Fairbrothers, 2006). Munther and Fairbrothers (2006) discovered that fern

spores treated with sporophyte leaf extract did not divide, and that fronds of Thelypteris normalis inhibited gametophyte growth of T. normalis, Pteris, and Phlebodium. Additionally, frond leachates from Pteridium and Dennstaedtia inhibited germination in Prunus, Rubus, Pseudosuga, Bromus, Hordeum, and Avena (Stewart, 1975; Star, 1980; Munther and Fairbrothers, 2006). Toxic inhibitors from fern fronds have the ability to disrupt development early in the gametophyte generation. These studies (Stewart, 1975; Star, 1980; Munther and Fairbrothers, 2006) show that a variety of fern species are capable of both producing allelopathic compounds, and displaying sensitivity to allelocompounds from ferns, and results from this study suggest that terrestrial ferns may also display sensitivity to epiphytic bryophytes. An allelopathic effect of epiphytic bryophytes on terrestrial gametophyte development seems evident. Because allelopathic compounds have been documented in bryophytes, and gametophytes are sensitive to various allelochemicals, it is hypothesized that the irregularities in development of terrestrial gametophytes observed in this study, are the result of such allelopathy. Abnormalities in gametophyte growth only occurred in terrestrial species that were sown directly on, or in proximity to, the epiphytic bryophytes Lejeuna flava and Calymperes palisotii. It is likely that terrestrial gametophyte establishment in epiphytic habitats is at least partially (if not largely) constrained by the presence of alleochemicals of epiphytic bryophytes, and possibly other epiphytic organisms as well. Epiphytic gametophytes were unaffected by the presence of bryophytes, implying that epiphytic species are resistant to these allelopathic compounds. The presence of these allelocompounds may in fact help reduce competition for space in epiphytic habitats. Because ferns and bryophytes share such a finite niche in epiphytic habitats, it is possible that ferns might reap benefits from the antibiotic, antibacterial, and antifungal properties that are prevalent in bryophytes by cohabiting with them. Additionally, fertilization in fern gametophytes is probably

facilitated by the presence of bryophytes because of their excellent ability to hold water, thus assisting with the movement of sperm. The current study identifies a feasible mechanism through which terrestrial ferns are excluded from epiphytic habitats. However, it does not address why or how epiphytic ferns do not grow in terrestrial habitats. Epiphytic ferns in terrestrial habitats may be less competitive for space and nutrients, or perhaps they are consumed by microherbivores. Additional research in this area would contribute to a more complete understanding of species establishment and habitat restrictions in epiphytic and terrestrial fern communities.

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Size (mm)

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Curriculum Vitae Mirabai Rachel McCarthy 316 Pearson Hall, Department of Botany, Miami University, Oxford, OH 45056 Phone: (802) –274-1124 email: [email protected]

EDUCATION

Graduate School (current) – Miami University, Oxford, Ohio 45056 Major - Botany Degree Sought – Doctor of Philosophy in 2011

Graduate School – Miami University, Oxford, Ohio 45056 Major - Botany Degree – Masters of Science, 2007

College - Lyndon State College, Lyndonville, Vermont Major – Environmental Science Minors - Biology and Geology Degree – Bachelor of Science, May, 2002

EDUCATOR EXPERIENCE

Graduate Teaching Assistant. Department of Botany, Miami University, Oxford, Ohio. August, 2005 - May, 2007. Courses: General Botany (BOT191) Environmental Education (BOT351) Dendrology (BOT205) Evolution of Plant Biodiversity (BOT204)

Instructor for Summer Workshop. Department of Botany, Miami University, Oxford, Ohio. Summer, 2006. Course: Incorporating Botany into Elementary and High School Science

Instructor. The King George School, Sutton, Vermont. February, 2003 – August, 2005. Courses: General Biology Algebra

Instructor. Lyndon State College, Lyndonville, Vermont. February – March, 2003. Course: Plant Growth and Function (BIO214)

Tutor. Lyndon State College, Lyndonville Vermont. December, 2001-May, 2003. Courses: Plant Growth and Function (BIO214) Plant Development (BIO213) Principles of Chemistry (CHE215)

Instructor. North Woods Stewardship Center, E. Charleston Vermont. Americorps Volunteer. August, 2002 – September, 2003 Workshops/Courses: Field Botany Environmental Education/Ecology Animal Tracking, Navigation, Rock Climbing, Canoeing, & Mt Biking

RESEARCH EXPERIENCE

Current: Morphology and Systematics of the Fern Genus, Adiantum. Research conducted under Dr. R. James Hickey. Department of Botany, Miami University, Oxford, Ohio.

2005 – 2007: Bryophyte Influence on Terrestrial and Epiphytic Fern Gametophytes. Research conducted under Dr. R. James Hickey. Department of Botany, Miami University, Oxford, Ohio.

2003: Plant, Insect and Fish Identification, and Mapping of the Connecticut River Islands. Research conducted under Dr. Dave Conant. Department of Biology, Lyndon State College, Lyndonville, Vermont.

2002 – 2003: A Comparative Spore Bank Analysis of Two Fern Populations; Athyrium filix-femina, and Dryopteris intermedia. Research conducted under Dr. Dave Conant. Department of Biology, Lyndon State College, Lyndonville, Vermont.

2001 – 2002: Potable Water Sources For Under Developed Communities in Honduras and El Salvador. Research conducted under the Monteverde Cultural Exchange, and Dr. Allison Lathrop. Department of Biology, Lyndon State College, Lyndonville, Vermont.

2000: Mycorrhizal Associations with Pteridophytes in the Panicum Cloud Forest, Puerto Rico. Research conducted under Dr. Dave Conant. Department of Biology, Lyndon State College, Lyndonville, Vermont.

FIELD EXPERIENCE

Bahamas, North Andros Island, 2006. Department of Botany, Miami University, Oxford, Ohio. Focus: Plant systematics, taxonomy, and island ecology.

Costa Rica, 2003 & 2001. Department of Botany, University of Vermont, Burlington,Vermont, and Department of Biology, Lyndon State College, Lyndonville,Vermont. Focus: Tropical plant systematics, taxonomy, and ecology.

Florida & Georgia, 2002. Department of Biology, Lyndon State College, Lyndonville,Vermont. Focus: Taxonomy of aquatic plants, and grasses.

Puerto Rico, 2000. Department of Biology, Lyndon State College, Lyndonville, Vermont. Focus: Plant systematics, taxonomy, tropical ecology.

ADDITIONAL WORK EXPERIENCE

Herbarium Assistant Miami University, Oxford, OH Aug, 2007 – current

Ecologist North Woods Stewardship Center, East Charleston, Vermont 2003 - 2004

Herbarium Assistant Fairbanks Museum, St Johnsbury, Vermont March – June, 2003

Creel Surveyor Vermont Department of Fish & Wildlife, St Johnsbury, Vermont May – December, 2003

Art Gallery Owner The Rising Sun – Global Artworks, Madrid, New Mexico May, 1996-April, 1997

PRESENTATIONS

* Thesis Defense, Miami University, Oxford, OH. September, 2007 Bryophyte Influence on Terrestrial and Epiphytic Fern Gametophytes * ASPB annual meeting, Chicago, Illinois, summer, 2007 Bryophyte Influence on Terrestrial and Epiphytic Fern Gametophytes * Plant Physiology Course, Miami University, Oxford, OH. Spring, 2006. Comparing the Rates of Photosynthesis Between Gametophytes and Sporophytes * Plant Development Course, Miami University, Oxford, OH. Fall, 2005. Antheridiogen Production and Response in Polypodiaceae Species. * Commencement Speaker, Lyndon State College, VT. Spring, 2002. Commencement address

MEMBERSHIPS/HONORS American Fern Society, 2007 Member Sigma Zeta Honorary Math/Science Society, 2003 Member Alpha Sigma Lambda National Adult Learner Society, 2002 - 03 Vice President, Natural Science Society L.S.C. 2000-02.

GRANTS/SCHOLARSHIPS Academic Challenge, Start-Up Grant, $250.00 Academic Challenge Grant, $750.00 W. Hardy Eshbaugh and Thomas K. Wilson Scholarship in Botany, $300.00

SKILLS * American Red Cross Life Guard, First Aid, and CPR for the Professional Rescuer, certification, 2004 * Wilderness First Responder, certification, 2003 * Field navigation using map & compass orienteering, & aerial photos * ArcView GIS, GPS Garmin