Mandrillus Leucophaeus Poensis)

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Ecology and Behavior of the Bioko Island Drill (Mandrillus leucophaeus poensis)

A Thesis

Submitted to the Faculty

Drexel University

Jacob Robert Owens

in partial fulfillment of the

requirements for the degree

Doctor of Philosophy

December 2013

Dedications

To my wife, Jen.

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Acknowledgments

The research presented herein was made possible by the financial support provided by Primate Conservation Inc., ExxonMobil Foundation, Mobil Equatorial Guinea, Inc., Margo Marsh Biodiversity Fund, and the Los Angeles Zoo. I would also like to express my gratitude to Dr. Teck-Kah Lim and the Drexel University Office of Graduate Studies for the Dissertation Fellowship and the invaluable time it provided me during the writing process. I thank the Government of Equatorial Guinea, the Ministry of Fisheries and the Environment, Ministry of Information, Press, and Radio, and the Ministry of Culture and Tourism for the opportunity to work and live in one of the most beautiful and unique places in the world. I am grateful to the faculty and staff of the National University of Equatorial Guinea who helped me navigate the geographic and bureaucratic landscape of Bioko Island. I would especially like to thank Jose Manuel Esara Echube, Claudio Posa Bohome, Maximilliano Fero Meñe, Eusebio Ondo Nguema, and Mariano Obama Bibang. The journey to my Ph.D. has been considerably more taxing than I expected, and I would not have been able to complete it without the assistance of an expansive list of people. I would like to thank all of you who have helped me through this process, many of whom I lack the space to do so specifically here. However, I would like to express my gratitude to: My advisor, Gail Hearn, for convincing me to switch to primates, and providing me with what has been the most exciting and rewarding venture of my life. Words cannot express how grateful I am for the opportunity you provided me to study one of the most interesting and least known primates, in one of the most fascinating locations. Thank you for letting me find my own way, for giving me the independence to make mistakes, and providing me the support to learn from them. I am so much more capable as a result. Shaya Honarvar, who has filled a large number of roles over these years, each as important to me as the next. As a committee member, she worked tirelessly to help develop and refine this project, and edit countless reports, proposals, presentations, posters, and chapters. In the field she provided the logistical support, data, volunteers, and samples from Playa Moaba. As a friend, she gave me the support, encouragement,

and reality checks I needed to continue. If it was not for our walks, I would not be writing this acknowledgement today. Michael O’Connor, for his uncanny ability to make me continually question what I think I know, and for making me strive to answer that question. His guidance has helped to build my self confidence in statistical analysis and hypothesis testing, and ultimately in my ability to succeed in this field. Susan Kilham, for her positivity, and for always helping me to see the bigger picture. Your insight has helped me to appreciate the interrelatedness of the world, and think more broadly about the theory and impact of ecology and evolution. Katy Gonder, for her consistently upbeat personality, excitement, and interest. Her knowledge of the ecology and evolution of primates helped me to shape this study into a coherent, manageable, and worthwhile endeavor. Christos Astaras, for the camaraderie, and for sharing with me your knowledge of drills, data and photographs, and methodological tips and tricks. Richard Bergl, Nelson Ting, and Josh Linder, for their technical, theoretical, and field assistance, and for introducing Drew and me to the primatology community with only a moderate amount of hazing. All those involved with the BBPP and Drexel Study Abroad, including Sally Vickland, Elizabeth Congdon, Heidi Ruffler, and Daniela Ascarelli. David Montgomery, Mark Andrews, Andrew Fertig, Karim Shnan, Matt Cuzzocreo, Livy Lewis, Nabil Nasseri for the logistical support, friendship, and much needed good times while in Bioko. Mary and Pete Johnson, for always treating me like family and for the best sausage gravy and biscuits in the eastern hemisphere. All of the Equatoguinean field workers who continually impress me with their skill and knowledge in the field. I would especially like to thank Cirilo Riaco, Miguel Angel Silochi, Fermin Muatiché, and Florentino Motove for collecting fecal samples, and for laughing at my excitement about said fecal samples. Filemon Rioso Etingue, the single most capable person I have ever had the opportunity to work with in the field. Thank you for teaching me how to identify Bioko’s primates and how to work and live in tropical Africa, and also for the friendship.

Esteban Muatiche, for opening your home to me, your friendship, work ethic, and beautifully high falsetto. I will always cherish our daily breakfasts and nightcaps together. My field assistants Brent Barry, Krissy Copeland, Tess Dornfeld, Julianna Gehant, Elissa Gordon, and Colleen Weathers, and lab assistants Rumaan Malhotra, Gian Bonetti, Atika Mehmood, Mark Nessel, Tyler Short, and Christian Brown, who volunteered a portion of their lives to drill feces and made this dissertation possible. Justin Jay, for countless hours in the blind watching drills from a few meters away. Thank you for the conversations, laughs, support, and thank you for your effort to help conserve the drills. Stephen (Steve Dubbs) Woloszynek, for your help with stats, conversations about religion and politics, and the inane video forwards at 4 am. Faculty and staff of the Drexel University Bio/BEES departments, especially Susan Cole, Ken Lacovara, Dan Duran, and Walt Bien. My fellow travelers of the Drexel University graduate experience. I was initially drawn to Drexel University because of the camaraderie and honesty I saw between the graduate students in this department. Over these years, I have come to realize that I had stumbled into something unique and extremely special in the world of academia. The friendship and support we have shared has enriched not only my graduate work, but also my outside life. Thank you all so much. The Hearn Lab members, for everything. Drew, Pat, Deme, and Miki, you have made life and field work so much more enjoyable. Thank you for all of the help and for the amazing and often ridiculous memories, working with you guys has been unforgettable. My friends from CHS and Stockton who continually stick by me, support me, and never take me seriously. Daniel Duran, Ilene Bean Eberly, Margaret Lewis, Ekaterina Sedia, Linda Smith, Roger Wood, and the other faculty of Richard Stockton College who convinced me I wasn’t finished with a bachelor’s degree, and have continued to support me ever since. Coach Todd Curll, Matt Markey, Bill Pino, Matt Waterhouse, Kim Kryscnski, Jason Mackie, and the other vaulters at RSC for giving me sporadic breaks from my

terrestrial existence. Lily, Tuba, Noodle, and Oscar, for their love, the excuse to take a break and go outside, and for helping me maintain my sanity during this process. I would like to thank my family for their enduring support and belief in me. There is no way I can express how truly grateful I am for all of your patience, love, reprieve, and food. The Jones and Edwards family, thank you for the continual support and for taking me in as family. Levi and Sarah, you are the best siblings I could have ever asked for, thank you for your patience, friendship, and the amazing nieces and nephews who have brought me so much joy. Mom and dad, you made this possible through your encouragement, love, support, and by always trusting me to make the right decisions in my life. Thank you for investing in my future, and for giving me the opportunity to see the world. Finally, my wife and best friend, Jen, you are my foundation. You are the most selfless person I know, and I am so incredibly lucky to have been with you all of these years. Thank you for always telling me to pursue my aspirations, and working to making sure I achieve them. I am eternally grateful.

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Table of Contents

LIST OF TABLES ...... x

LIST OF FIGURES ...... xii

ABSTRACT ...... xiv

CHAPTER 1: INTRODUCTION TO THE DISSERTATION ...... 1 Introduction to the Drill ...... 1 Physical Description and Phylogeny ...... 1 Range ...... 4 Conservation Status ...... 5 Previous Studies ...... 9 Dissertation Objectives ...... 10 Description of the Study Sites ...... 11 Primate Community ...... 13 Field Sites ...... 14

CHAPTER 2: DIET AND FEEDING ECOLOGY ...... 21 Introduction ...... 21 Methods ...... 27 Study Site ...... 27 Food Availability Estimates ...... 28 Fecal Sample Collection and Analysis ...... 29 Data Analysis ...... 31 Ethical Note ...... 33 Results ...... 34 Food Availability Estimates ...... 34 Qualitative Dietary Analysis ...... 34 Quantitative Dietary Analysis ...... 37 Discussion...... 40 Diet and Conservation ...... 45 Conclusions ...... 46

CHAPTER 3: GASTROINTESTINAL PARASITIC INFECTIONS ...... 58 Introduction ...... 58 Methods ...... 65 Study Site ...... 65 Subjects ...... 66 Sample Collection ...... 67 Data Analysis ...... 68 Results ...... 69 Parasite Prevalences by Site ...... 69 Parasitic Infections and Diet ...... 69

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Discussion...... 70 Parasites of the Papionin ...... 74 Implications for Drill Conservation and Human Health ...... 78 Conclusions ...... 79

CHAPTER 4: GROUP SIZE, POLYSPECIFIC ASSOCIATIONS, AND HABITAT USE ...... 84 Introduction ...... 84 Methods ...... 90 Study Design and Caveats ...... 90 Group Size, Composition, and Polyspecific Associations ...... 91 Habitat Profile ...... 92 Data Analysis ...... 94 Results ...... 95 Group Size, Composition, and Associations ...... 95 Habitat Assessments ...... 97 Discussion...... 99 Group Size and Associations ...... 99 Habitat use ...... 107 Conclusions ...... 109

CHAPTER 5: STABLE ISOTOPE ECOLOGY ...... 114 Introduction ...... 114 Methods ...... 122 Ethical Note ...... 122 Sample Collection and Preparation ...... 122 Data Analysis ...... 125 Results ...... 127 Food Sources ...... 127 Consumer Isotopic Variations ...... 129 Dietary Source Proportions ...... 130 Discussion...... 132 Consumer Values ...... 133 Sexual Differences ...... 137 Seasonal Differences ...... 138 Niche Overlap and Competition ...... 139 Carcass Origins ...... 140 Conclusions ...... 142

CHAPTER 6: DISSERTATION CONCLUSIONS AND BROADER IMPLICATIONS ...... 155 Synopsis of the Dissertation ...... 155 Feeding Ecology ...... 156 Gastrointestinal Parasitic Infections ...... 157 Group Size, Polyspecific Associations, and Habitat Use ...... 158 Stable Isotope Ecology ...... 159

Limitations and Future Directions ...... 160 Conservation Implications ...... 162

LIST OF REFERENCES ...... 164

VITA...... 192

List of Tables

Table 2.1: Fecal sample collection effort and results for Bioko Island drills at the three study sites ...... 50

Table 2.2: List of vegetative food items consumed by Bioko Island drills in this study ...... 51

Table 2.3: Comparison of the prevalence, species richness, diversity, and evenness of seeds and invertebrates within the fecal samples at each field site ...... 53

Table 2.4: List of animal food items consumed by Bioko Island drills in this study .....54

Table 2.6: Principal component scores of the dietary category masses ...... 56

Table 3.1: Taxonomy, pathology, and prevalence of the parasitic infections of the Bioko Island drill within the Caldera (montane forest) and Moaba Playa (lowland forest). The Strongyle eggs were likely from Oesophagostomum sp., however confirmation was not possible due to the absence of adult worms ...... 83

Table 4.2: The proportion (%) of polyspecific associations recorded during encounters with Bioko Island drills (N = 143) and drills in Korup N.P., Cameroon (N = 44), as reported by Astaras et al. (2011) ...... 112 Table 5.1: Summary results for the analysis of food sources, including the total nitrogen and carbon of the tissues and their stable nitrogen and carbon values ...... 151

Table 5.2: Results of the ANOVA's and subsequent multiple comparisons (Tukey's Post Hoc Test, p value) of the δ13C‰ (top) and δ15N‰ (bottom) values of food sources ...152

Table 5.3: Summary information for the stable isotope analysis of consumer hair samples ...... 153

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List of Figures

Figure 1.1: Molecular phylogeny and estimated divergence times of the African Papionin monkeys (Tosi et al. 2003) ...... 16

Figure 1.2: The Papio of Genser (1554, p 15) referenced by Linnaeus (1758) in the type description of Papio (Mandrillus) sphinx, the mandrill...... 17

Figure 1.3: Full extent of the current range of the drills in the Cross-Sanaga-Bioko rainforests as recognized by the IUCN Red List (IUCN 2013) ...... 18

Figure 1.4: Locations of the two recognized protected areas, major roads, and the capital city, Malabo, on Bioko Island (left). Expanded view of the GCSH, showing the locations of the three study areas (Caldera, Moraka Playa, and Moaba Playa), transects, and the primary forest types in the area (right) ...... 19

Figure 1.5: Variation in the average monthly rainfall of two villages, Moka and Ureca, in montane and lowland forest, respectively, in southern Bioko Island. Figure adapted from Font Tullot (1951) ...... 20

Figure 2.1: Mean weight (left) and volume (right) of the dietary categories by their absolute values (top) and proportion within the total diet (bottom) ...... 47

Figure 2.2: Bivariate plot of the first two principal components scores for each location ...... 48

Figure 2.3: Pairwise comparison of the dietary overlap between the sites, as indicated by Czekanowski’s proportional similarity index (PS) ...... 49

Figure 3.1: Counts of the species richness of parasites found in drill fecal samples collected at the Caldera and Moaba Playa ...... 81

Figure 3.2: Frequency of parasitic infections occurring in fecal samples collected at the Caldera and Moaba Playa ...... 82

Figure 4.1: Group composition based on reliable encounters from transects performed at all locations and blinds set at Moaba Playa (N given in legend, error bars = SEM). Asterisks denote a significant difference between the survey method estimates ...... 110

13 15 Figure 5.1: Boxplots of δ C and δ N isotopes (top) and total %C and %N (bottom) for C3 plants, C4 grasses, and invertebrate food sources ...... 144

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13 Figure 5.2: Relationship between the carbon isotope values (δ C‰) of herbaceous plant tissues and elevation ...... 145

13 15 Figure 5.3: Boxplots of the δ C‰ (left) and δ N‰ (right) values of consumer hair samples ...... 146

13 Figure 5.4: Minimum convex polygons of each species and their sex classes in the δ C 15 and δ N niche space. Total area occupied (“δ space”) by the polygons and nearest neighbor values (mean +SEM) are provided ...... 147

13 15 Figure 5.5: Plots of the δ C and δ N values for each species. Points correspond to each individual sampled with their sex class denoted by color; those with error bars (SD) represent the means. Kernel density estimates of the points are shown as contour lines (middle plots) for the niche space biplots, and separately for the corresponding frequency 13 15 distributions of the δ C and δ N (bordering plots) ...... 148

Figure 5.6: Stable isotope values of consumer hair (symbols: individual means corrected 13 15 by the TEF values) and source tissues (symbols: mean + SD) plotted in the δ C - δ N niche space ...... 149

Figure 5.7: SIAR boxplots of the predicted proportions of each food source to the diets of the consumer species (boxes represent the 95, 75, and 25% credibility intervals) .....150

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ABSTRACT

Ecology and Behavior of the Bioko Island Drill (Mandrillus leucophaeus poensis) Jacob Robert Owens Advisor: Gail W. Hearn, Ph.D.

Despite once ranging across Equatorial Guinea’s Bioko Island, drill monkeys

(Mandrillus leucophaeus poensis) are now limited by intense bushmeat market hunting to

the Gran Caldera and Southern Highlands Scientific Reserve, a nominally protected area

that comprises the southern third of the island (550 km2). Few studies have investigated

the ecology or behavior of wild drills, and none have been performed on the endemic subspecies on Bioko Island. The objective of this dissertation was to provide a robust understanding of several fundamental aspects of the natural history of the Bioko drill.

Comparison of foraging observations and fecal samples collected between montane and lowland forest habitats during the dry seasons of three consecutive years (2009-2011) revealed distinct dietary patterns within each habitat. In lowland forests, where fruits were relatively abundant, drills were primarily frugivorous, with only 10% of their fecal

samples composed of non-fruit food items. In the montane forests, where fruits were

scarce, their diet was primarily composed of the pith of terrestrial herbaceous vegetation.

This fallback diet was also marked by significant increases in the weight and volume of fecal samples, and in the consumption of insects, leaves, and mushrooms. Analysis the gastrointestinal parasites of Bioko drills found them to be infected by at least six parasite species common to other primates. This is the first study to report the coccidian species,

Cyclospora papionis, outside of olive baboons (Papio anubis) in east Africa, representing a considerable expansion of its range. Reliable estimates of drill group sizes were made during 136 encounters. Mean group size was 3.8 individuals per group (SEM = + 0.3;

range = 1-20), and on average, groups contained one adult male and one female. Stable

isotope analysis of hair samples collected from drills and other medium sized mammals

indicated that the dietary overlap between the primate and duiker species may be high.

However, there was little evidence that this potential competition was resulting in niche

shifts among species. The findings of this research contrast some of the longstanding

assumptions of the ecology and behavior of the drill, and provide important information

for the conservation of the species.

CHAPTER 1: INTRODUCTION TO THE DISSERTATION

Introduction to the Drill

Physical Description and Phylogeny

Drills (Mandrillus leucophaeus F. Cuvier, 1807) are a large bodied, highly sexually dimorphic primate species found within the lower Guinean forests of Central

Africa. There are currently two recognized drill subspecies. The mainland drill (M. l. leucophaeus, Cuvier, 1807) is found in a range that spans from the Sanaga River in

Cameroon to the Cross River in Nigeria, while the Bioko drill (M. l. poensis, Zukowski,

1922) is endemic to Bioko Island, Equatorial Guinea. Bodyweight varies between the two

subspecies, with the means of adult Bioko drills (male: 20.0 kg; female: 8.5 kg) less than

the mainland drills (male: 32.9 kg; female: 11.6 kg) (Butynski et al. 2009, 2013). This

discrepancy is largest between males, resulting in greater sexual dimorphism in the

mainland drill.

Pelage is less variable between populations, but differs dramatically between

adult males and other sex/age classes. Both males and females are predominantly

greenish grey to olive-brown, with grey-white ventrum, inner limbs, and chinstraps.

Faces are black and hairless, with elongated muzzles and large, boney paranasal ridges.

Adult males are distinguished by large medial crest, longer shoulder and upper back hair

(forming a mane), a bright white chinstrap, and longer, greyish hairs mid-chest

surrounding the sternal gland (Butynski et al. 2013). Skin coloration in adult males

includes a pink/scarlet red patch below the bottom lip, deep red in the inside of thighs,

groin, and red-blue anogenital regions (Hill 1970). The paranasal ridges, cheek flanges, 2

and canines of males are considerably larger than females. These secondary sexual

characteristics vary between males, being most dramatic in dominant males (Marty et al.

2009).

Drills are an Old World monkey (Family: Cercopithecidae; Subfamily:

Cercopithecinae) within Papionin tribe, which consists of the drill and their congener, the

mandrill (Mandrillus), mangabeys (Cercocebus spp. and Lophocebus spp.), macaques

(Macaca spp.), baboons (Papio spp.), gelada (Theropithecus gelada), and the recently discovered Kipunji monkey (Rungwecebus kipunji) (Grubb et al. 2003; Jones et al. 2005).

The taxonomy and phylogeny of the African Papionins have been heavily debated in the

200 years since their discovery by western scientists (Fleagle & McGraw 2002). For much of this time they were placed into two groups based on conspicuous morphological characteristics, including Cercocebus (mangabeys) and Papio, the latter consisting of savannah baboons, gelada, and drills and mandrills, then referred to as the “forest baboons” (Fleagle & McGraw 1999). More recent evidence from increasingly sophisticated molecular analyses (e.g. Disotell et al., 1992; Disotell, 1994, 1996; Harris &

Disotell, 1998; Olson et al., 2008; Burrell et al., 2009) and more extensive osteological comparisons (e.g. Fleagle & McGraw, 1999, 2002; Gilbert, 2007; Gilbert et al., 2009) have consistently placed Mandrillus and Cercocebus in a monophyletic group, however the relationship between the other Papionin genera is less resolved. Molecular dating techniques estimate the divergence of the Mandrillus/Cercocebus clade from the other

Papionins at 9.0 to 11.5 mya, and the subsequent split of the two genera at 3.2 to 4.1 mya

(Harris 2000; Tosi et al. 2003) (Figure 1.1).

This historic difficulty in elucidating the phylogeny and taxonomy of drill was not

limited to their relationships with the other Papionin genera, as numerous discrepancies

exist in the literature concerning the Mandrillus nomenclature since their discovery

(Delson & Napier 1976). Even the type description is questionable. In the first

description of the mandrill (Simia sphinx) by Linnaeus in Systema Naturae, vol 1 (1758,

p. 25), he provides the description, “short tailed monkey, facial vibrissae, pointed claws”,

for the mandrill, but makes no mention of the “blue furrowed cheeks”, a characteristic

unique to mandrill, that he later describes of Simia maimon (Linnaeus 1766). With his

description of Simia sphinx, Linnaeus references Gesner’s Historia animalium de

quadrupedibus viviparis (1554) 1, depicting a baboon-like monkey that Gesner saw on

exhibition in Augsburg, Germany, in 1551. The animal, which Gesner refers to as

Arctopithecum (“bear-ape”) and Cynocephalus (“dog-head”), has relatively large head,

neck, and shoulders, long whiskers, short tail, and is completely dark in color (Figure

1.2). Given that the drawing is of a male, we should expect some description of the

unique coloration of its face or anogenital region, if it indeed depicted a mandrill. Based

on these and other discrepancies, Delson & Napier (1976) suggest that the initial species

description of the Simia sphinx, now attributed to the mandrill, may have actually been of

a drill. However, as the whereabouts of subject of Gesner’s drawing are unknown, there

is no way to confirm this suspicion.

Because of this confusion, at least seven and 14 species synonyms exist for drills

and mandrills, respectively (Wilson & Reeder 2005), and both have been placed in at

least six genera, including Papio and Simia, before being moved to Mandrillus (Ritgen,

1 Linnaeus incorrectly cited pages 352‐353, however it is now attributed to pg. 15 of the appendix (Delson & Napier 1976).

1824) by Allen (1939). Currently there is a strong consensus on the two recognized

Mandrillus species (M. leucophaeus and M. sphinx). However, there is less evidence to support the distinction of the drill subspecies (Grubb et al. 2003).

Range

Drills are endemic to the Cross-Sanaga-Bioko rainforest region of western Central

Africa. The historic range occupied is estimated at less than 50,000 km2, thought to be

one of the smallest of all the medium to large African mammals (Gadsby & Jenkins 1997;

Eeley & Foley 1999; Butynski et al. 2013). Prior to 1987, drills were thought to be

extirpated in both Nigeria and on Bioko Island, and the only confirmed population

remaining was in Korup National Park, in North-Western Cameroon (Oates 1986; Gadsby

& Jenkins 1997). However, subsequent surveys performed in the late 1980s confirmed

their presence in these areas (Lee et al. 1988; Butynski & Koster 1994).

On the mainland, drills range from the Cross river in Nigeria to the Sanaga River

in Cameroon (Figure 1.3). Roughly 80% of the species range exists in Cameroon, spanning 20,000 km2 in at least nine fragmented forest blocks (Gadsby & Jenkins 1997;

Wild et al. 2005). Recent surveys of the country wide range in Cameroon by Morgan et

al. (2013) suggest that they are likely extirpated from 46% (24/52) of the survey units

previously occupied by drills. These surveys also indicate that the largest populations are

currently found in and around Korup National Park and Ebo Forest, in the northwest and

southeast extents of their Cameroonian range, respectively (Morgan et al. 2013).

Surveys on Bioko Island between 1986 and 1994 reported drills within two

fragmented forest units of a combined area less than 800 km2 (Butynski & Koster 1994;

Gonzalez-Kirchner & de la Maza 1996) (Figure1.3). Each unit is in one of the two

nationally recognized protected areas on the island, Pico Basilé National Park and the

Gran Caldera and Southern Highlands Scientific Reserve (GCSH). Within the GCSH, which covers roughly the southern third of Bioko Island, drills are found from the coastal beaches to approximately 1000m asl (Butynski & Koster 1994; Gonzalez-Kirchner & de la Maza 1996). The relative abundance of drill groups at survey sites in the GCSH (0.1

groups/km) is the highest of any location in their range, and an order of magnitude higher

than that of Korup National Park (0.01 groups/km) (Astaras et al. 2008; Linder & Oates

2011; Cronin 2013).

The range of drills on Pico Basilé N.P. is relatively unknown, but is thought to be

restricted to the southern slopes between 600 to 800m asl (Butynski & Koster 1994). The

last reported encounters on Pico Basilé were from surverys performed in 1986 by

Butynski & Koster (1994) and in 1989-1990 by Gonzalez-Kirchner & de la Maza (1996).

Butynski & Koster found drills at only one location within the Pico Basilé range, and

Gonzalez-Kirchner & de la Maza note that a small population existed near the village of

Moeri. Subsequent surveys on Pico Basilé, including the areas surrounding Moeri, have

not encountered drills in this area (Butynski & Owens 2008; Vega et al. 2013), and ad

hoc discussions with hunters in Moeri indicate that they may now be extirpated (pers.

obs.).

Conservation Status

The drill is listed as an IUCN Endangered species and CITES: Appendix I species,

primarily because of the intense and illicit commercial bushmeat hunting activities and

habitat loss that exist throughout their range (Gadsby & Jenkins 1997; Wild et al. 2005;

Morgan et al. 2013).

Drills are a preferred bushmeat, and their large body size and high cost-to-weight ratio

increases the cost profitability to hunters, leading to them being disproportionately

targeted during hunting activities compared to most primates (Reid et al. 2005; Astaras

2009). Long-term bushmeat surveys performed at Semu bushmeat market in Malabo,

Bioko Island, have shown that over a ten year period (1997-2007), drills contributed 8.2%

of the total bushmeat biomass of the market, higher than any other primate (Cronin 2013).

These surveys also show that the rate at which drill carcasses sold each day has increased

significantly since 1997, and as of 2010, roughly five drill carcasses were being sold each

day the market was open (six days per week) (Cronin et al. 2010).

The use of dogs in shotgun hunting has been recognized as the primary threat to

the continued existence of drills throughout most of their range (Butynski & Koster,

1994; Gadsby et al., 1994; Gadsby & Jenkins, 1997; Steiner et al., 2003; Wild et al.,

2005; Astaras, 2009; Morgan et al., 2013). Drills are not as adept at arboreal movement as

the other monkeys throughout their range, and with the use of hunting dogs, hunters can

force entire groups into trees where they can shoot a large proportion of group members

at one time (Colell et al. 1994; Gadsby & Jenkins 1997; Astaras 2009). In Korup N.P.,

Cameroon, Steiner et al. (2003)found that 97% of hunters surveyed (N = 42) said they

used dogs to hunt drills, and the male was always targeted first, primarily because of its

high value. In the village of Moka, Bioko Island, Colell et al. (1994) found that dogs were

used in 38% of hunts, and the success rate of shotgun hunting was highest in drills among

all primates. This was reflective of the consensus among all hunters surveyed, that dogs

were essential to the success of hunting drills (Colell et al. 1994).

All three countries in the range of drills have laws or decrees banning the take of endangered species. In Cameroon, Ministerial order No. 0565 bans all hunting of drills,

and carries fines can be as high as 10,000,000 CFA for each individual monkey carcass,

as well as potential imprisonment (Wild et al. 2005). In Nigeria, the Endangered species

act of 1985 (Decree No. 11) lists the drill as a Schedule 1 species, and has various fees associated with any capture, hunting, or trade. In Equatorial Guinea, a presidential decree

(Ley num. 72/2007) bans the hunting, sale, or consumption of primates, carrying fines of

100,000 – 500,000 CFA per individual monkey, and potential imprisonment (EG 2007).

However a lack of enforcement by the governments of any of these countries has resulted

in a continual deterioration of the conservation status of the drill, and additional local

extirpations are likely to occur.

Habitat fragmentation and degradation is not currently a major threat to the

population on Bioko Island, and there is indication that a decreased use of agricultural and

pasture lands may be increasing the available habitat for primates on the island (Butynski

& Koster 1994). Much of the area where drills are currently found is characterized by

dramatic topographic variation, including steep ravines, cliffs, and craters, making it unlikely that much of this area will ever be developed. However, continued development on the mainland, particularly from the development of palm oil plantations throughout their already limited range, does pose a serious and immediate threat to mainland drills

(Morgan et al. 2013).

Several long-term programs of national and international non-governmental organizations (NGOs) have worked to conserve drills, and other primate and non-primate

taxa, throughout their range. In Cameroon, groups such as the Conservation and Ecology

of Drills in Cameroon (CRES, San Diego Zoo), Wildlife Conservation Society (WCS),

German Development Agency (DED), and World Wildlife Fund (WWF) have been

integral to the development or ongoing maintenance of several national parks in areas

occupied by drills (Astaras 2009). The Bioko Biodiversity Protection Program (BBPP), a

partnership between the National University of Equatorial Guinea, Malabo, EG, and

Drexel University has been the only stable conservation organization on the island since

before 1998. The BBPP performs year-round surveys on both the forests and bushmeat

markets on Bioko to monitor the impact of hunting activities on the island’s fauna.

Conservation related activities also include educational programs and awareness

campaigns, and ecotourism, the development of a long-term field research station, and

advocacy.

The main organization focused specifically on the conservation of the drill has

been the Pandrillus Foundation (Pandrillus), and their associated projects, the Drill

Rehabilitation & Breeding Center (DRBC), founded in 1991 in Calabar, Nigeria, and the

Limbe Wildlife Center, founded in 1993 in Limbe, Cameroon. Pandrillus is involved in

numerous conservation efforts, including research, education, and advocacy, however

their primary activities have involved the captive care and breeding of drill orphans. The

DRBC and Limbe Center recover and care for orphaned drills being kept as pets illegally

(typically a result of hunting), and house them in large enclosures of natural forested areas

(Wood 2007). The DRBC has been successful at breeding these individuals, as over 200

have been born since 1991, and it now has the largest captive population of drills in the

world (Wood 2007). Pandrillus has been particularly effective in garnering local public

support by providing jobs and economic support to the area, and there is indication that

some commercial logging activities near Afi Mountain may have been abandoned

because of local involvement (Ewak 2010).

Previous Studies

The drill has been the focus of surprisingly few ecological or behavioral studies,

despite its conservation status, taxonomic distinctiveness, and potential importance to the

forests in its range. Empirical information on the reproduction and social interactions of

drills is limited to those in semi-captive conditions, studied at the DRBC by Wood

(2007), and there is no available information from wild individuals. The early, seminal

work on the of wild drills in Cameroon by Gartlan and Struhsaker in the Bakundu Forest

Reserve, northeast of Mount Cameroon (Struhsaker 1969; Gartlan 1970; Gartlan &

Struhsaker 1972), was followed by a 36 year gap in the literature until the publication of

the doctoral thesis of Astaras (2009) and associated papers (Astaras et al. 2008, 2011;

Astaras 2009; Astaras & Waltert 2010). The literature on Bioko drills is even more

sparse, coming primarily from the descriptive field notes while collected during primate

surveys (Schaaf et al. 1990; Butynski & Koster 1994; Gonzalez-Kirchner & de la Maza

1996). No direct focal studies have been performed on the feeding, grouping, ranging or

any other ecological aspects of this subspecies.

Dissertation Objectives

Ecological and behavioral data is critical to effective conservation and management programs, particularly as the resources available for actions are often limited

(Gogol-Prokurat 2011). Prioritization of threats, species, and habitats is dependent on the

habitat use and distribution of species across a landscape, and this information is crucial

to the design and prioritization of protected areas (Lacher & Alho 2001; Papes 2007). It is

therefore imperative that ecological and behavioral research focus on identifying the

ability of species to adapt to ecological variations in their habitat, as well as the factors

that may promote or inhibit the use. With this in mind, my overarching objective in this

dissertation was to provide a robust understanding of several fundamental aspects of the

ecology and behavior of the Bioko Island drill. The primary research questions I address

are:

1. What is the diet of the Bioko Island drill, and what impact does resource

availability have in shaping it?

2. What are the intestinal parasitic infections of Bioko Island drills?

3. What is the size, composition, and habitat use of drill foraging groups?

4. What is the extent of the dietary niche overlap between the drill and other

medium-large mammals Bioko Island?

In chapters two, three, and four, I use the extensive variability in habitat structure

and food availability existing between montane and lowland forest types to address these

questions. Each of these chapters also compare with what is known of the better studied

mainland drill and mandrill to broaden the context of the results. Chapter two (Diet and

Feeding Ecology) details the qualitative and quantitative dietary characteristics of the drill

based on field observations and fecal sample analysis to determine what strategies they

use to compensate for fruit scarcity. Chapter three (Gastrointestinal Parasitic Infections) investigates the variations in the species richness and prevalence of the gastrointestinal parasites of drills, and determine if environmental variations or dietary composition have

an impact. In chapter four (Group Size, Polyspecific Associations, and Habitat Use), I use

two survey methods, transects and blinds, to estimate the group size and composition and

polyspecific associations of Bioko Island drills, and determine the effect of the survey

method on the results. I also perform a habitat assessment to determine if there are

relationships between the location that drill groups are encountered and the habitat

features at a location. In chapter five (Stable Isotope Ecology) I use stable isotope

analysis of hair samples collected from bushmeat carcasses to assess the potential

interspecific and intraspecific competition of drills. Chapter six (Dissertation Conclusions

and Broader Implications) provides a synthesis of the information learned from the four

research chapters, discusses the limitations of this work and the future research

opportunities, and puts these findings in context to the conservation of the Bioko Island

drill.

Description of the Study Sites

Bioko Island, Equatorial Guinea (3.8 to 3.2 N and 8.4 to 8.9 E) is a steeply

volcanic, continental shelf island, separated from mainland Cameroon by a shallow, 37

km wide channel created by rising sea levels approximately 12,000 years ago. The

volcanic origins of the island have resulted in a topographically diverse landscape

including three volcanic peaks: Pico Basilé 3008 m asl. in the north and two smaller

peaks, Gran Caldera de Luba (2260 m asl.) and Pico Biao (2009 m asl.), in the south.

Bioko is divided into altitudinal rings of four distinct forest types (Butynski &

Koster 1989). The lowland rainforests (0-800m asl), are comprised of the most diverse

vegetative components with numerous Ficus spp., and other trees growing up to 40 m in

height (Juste and Perez del Val, 1995). Although much of the lowland forests were once

heavily degraded for logging or agricultural use, more than 570 km2 of pristine lowlands

still exist, making it the most widespread habitat on the island (Butynski & Koster 1994).

Montane forests (800 – 1400m asl), which have had relatively little anthropogenic

disruption, are characterized by a much lower canopy and vegetative diversity than the

Lowlands forests. Tree ferns (Cyanthea sp.) and an increase in epiphytes are typical of

this habitat. The Montane forest floors are densely covered by terrestrial herbaceous

vegetation, most notably Aframomum spp. and Bracken Fern (Pteridium aquilinum),

which form large colonies of dense fields. The Mossy forests (1400 – 2600 m) and

Shrublands (2600 – 3000 m) habitats, found only on Pico Basilé, are characterized by low

canopy heights, densely vegetated understories, and low plant diversity (Juste & Perez

Del Val 1995)(Juste and Perez del Val, 1995).

The mean annual temperature on Bioko Island is 25.1C, however, due to the

steep topography on Bioko Island, estimates are highly variable across the island (Terán

1962). The lowest estimated annual temperature on the island (8°C) is on the island’s

highest peak, Pico Basilé (3011 m), and highest at sea level on the northern coast

(26.5°C) (Font Tullot 1951). Temperature remains relatively stable throughout the year,

however latitudinal oscillations in the Intertropical Convergence Zone result in strong

seasonal rainfall variations (Oates et al. 2004). There is a single wet season that typically

begins in May, peaks in July-September, and ends in October (Nosti 1942; Font Tullot

1951) (Figure 1.5). Rainfall varies longitudinally across Bioko, with rainfall mean

average rainfall estimates of 1557 mm in the north, and 10934 mm on the southern coast

(Nosti 1942).

Primate Community

Bioko Island has been recognized as having high levels of species richness, but because of its close proximity and relatively recent separation from the mainland, the level of endemism of most taxonomic groups is low when compared to other islands in the Gulf of Guinea (Jones 1994). This is, however, true for the primate community on

the island, which is noted as having high richness and endemism, as well as a high

number of threatened taxa (Butynski et al. 2009). Within the 2017 km2 area of Bioko

there are seven species of monkeys from three genera, and four species of galagos from

three genera (Grubb et al. 2003; Butynski et al. 2009) (Table 1.1). The community

includes five species that are listed on the IUCN Redlist as vulnerable or higher,

including the Critically Endangered red colobus (Procolobus pennantii), and the

endangered Preuss’s monkey (Allochrocebus preussi) and drill. Seven subspecies are

recognized as endemics to Bioko Island, and all but one of these is listed as Endangered.

The monkeys fall into three general dietary guilds. Pennant’s red colobus

(Procolobus pennantii pennantii) and Bioko black colobus (Colobus satanas satanas) are

primarily folivorous-granivorous, the Bioko red-eared monkey (Cercopithecus erythrotis

erythrotis), Golden-bellied crowned monkey (Cercopithecus pogonias pogonias),

Stampfli’s putty-nosed monkey (Cercopithecus nictitans martini), and Preuss’s monkey

(Allochrocebus preussi) are arboreal frugivorous-omnivores, and the drill is primarily a

terrestrial frugivorous-omnivore. However, there is considerable overlap between the

species, and all are known to consume some amount of fruit (Butynski et al. 2013).

The average weight of the primates on Bioko Island varies from Demidoff’s dwarf galago, weighing less than 60 g on average, to the drills which reach a maximum weight of 45.0 kg (mean = 20.0) (Butynski et al. 2013). Male drills are substantially larger than all other mammals on Bioko Island. The average male weight is roughly 8 kg more than the next two heaviest species, the red duiker (Cephalophus ogilbyi) (mean of combined sexes = 11.9 kg) and the red colobus (male mean = 11.0 kg), and 14.5 kg more than the largest frugivorous monkey, Preussi’s guenon (20.0 kg vs. 5.5 kg) (Butynski et al., 2009).

Field Sites

Field work during this study was conducted in within the Gran Caldera and

Southern Highlands Reserve (GCSH) (Figure 1.4). The GCSH remains one of the least disturbed and most remote areas of the island, and the only remaining location where each of the island's seven native monkey species occur (Butynski & Koster 1994). There is only a single permanent human settlement within the GCSH, Ureca, a village of less than 200 residents. Although an access road has been in development for the past four years, currently the only way to travel to Ureca is a 20 km hike or a several hour boat ride.

Three field sites were used, including Moraka Playa and Moaba Playa, which

were within the coastal lowland forests at the extreme southwest and southeast sectors of

the GCSH, respectively (Figure 1.4). All samples and data collected from Moraka Playa

were from 300 m asl. or less, and most of them from Moaba Playa were less than 100m

asl., and within 1km of the beach line. The third site, Caldera, was located within the

volcanic crater of the Gran Caldera de Luba, in montane forest habitat from

approximately 900 – 1200 m asl. Patrols and transects at Moraka Playa and the Caldera

benefited from a series of transects established by the BBPP for primate surveys. In the

Caldera, the primary transect used was the Santo Antonio trail, a loop of 5.17 km in

length and roughly 1.8 km in diameter. Transects at Moraka Playa included the Tope

Tomo, Moraka Norte, and Badja South, and Badja North, a combined length of 12.8 km.

Numerous rivers were also regularly surveyed at all locations, and a considerable amount

of samples and data were collected off-trail. Moaba Playa is not a site of long-term

primate monitoring activities, and there are no established survey transects. Data was

collected entirely from rivers, coastlines, or in the forest on an ad hoc basis.

Figure 1.1: Molecular phylogeny and estimated divergence times of the African Papionin monkeys (Tosi et al. 2003).

Figure 1.2: The Papio of Genser (1554, p 15) referenced by Linnaeus (1758) in the type description of Papio (Mandrillus) sphinx, the

mandrill.

Figure 1.3: Full extent of the current range of the drills in the Cross-Sanaga- Bioko rainforests as recognized by the IUCN Red List (IUCN 2013).

Figure 1.5: Variation in the average monthly rainfall of two villages, Moka and Ureca, in montane and lowland forest, respectively, in southern Bioko Island. Figure adapted from Font Tullot (1951).

CHAPTER 2: DIET AND FEEDING ECOLOGY

Introduction

Approximately 90% of all primate species live in tropical rainforest regions

(Mittermeier 1988), which are often characterized by steep altitudinal gradients, dramatic

seasonal shifts, and highly heterogeneous landscapes. As a result, most primates are subject to high spatial and temporal fluctuations in the availability and distribution of

food resources (Hill 1997; Tutin et al. 1997). These resource variations have been shown

to influence numerous aspects of the ecology and behavior in a wide breadth of primate taxa, including dietary composition and diversity, social organization and behavior, ranging and habitat use, and reproductive timing (Brockman & van Schaik 2005).

For example, during periods of low fruit availability, grey-cheeked mangabeys

(Lophocebus albigena) in Dja Reserve, Cameroon, were found to consume significantly less fruit, and more seeds, flowers, and young leaves from a higher diversity of species

(Poulsen et al. 2001). This dietary shift corresponded with significant increases in the diversity of species consumed, time spent resting, and use of swamp habitats, while time feeding was significantly reduced and replaced with resting. Conversely, the relative consumption and diversity of fruit in red-capped mangabeys (Cercocebus torquatus) in

Campo Reserve, Cameroon, was relatively stable throughout the year (Mitani 1989).

Instead of switching to an alternative diet during periods of fruit scarcity, their core home ranges decreased and shifted to areas where preferred fruiting trees were in clumped distributions. Comparisons of Tana River mangabeys (Cercocebus galeritus) groups in eastern Kenya, found those in forests with low fruit availability to have decreased social

group sizes and fecundity, and increased daily travel distances and home ranges (Mbora

et al. 2009).

As demonstrated by these three closely related species, the ecological and

behavioral responses of primates to changes in food availability can be highly variable.

Comparative studies have shown that considerable differences exist in these responses

between sympatric species (Clutton-Brock 1974; Gautier-Hion 1980; Chapman 1987;

Vogel et al. 2009), neighboring conspecific populations and groups (Chapman & Fedigan

1990; Bronikowski & Altmann 1996; Chapman et al. 2002), and individual group

members (Post 1981; Boinski 1988; Barton & Whiten 1993; Isbell & Young 1993). For

instance, chimpanzee (Pan troglodytes) fruit consumption in Lopé Reserve, Gabon, was

higher within fragmented habitats, despite having relatively low fruit diversity and

availability, compared to continuous forests (Tutin 1999). The opposite was true for all

other species in the primate community. Population densities were higher in the fragmented habitats in half of the species, including the chimpanzees, mandrill

(Mandrillus sphinx), putty-nosed guenon (Cercopithecus nictitans), and moustached guenon (Cercopithecus cephus), densities of the latter being 23 times higher than in continuous habitats.

The high intraspecific variability that has been found in the ecology and behavior of primates has recently led some authors to warn against overgeneralizations of study findings and broad theoretical concepts (Ganas & Robbins 2004; Sayers 2013). Others have suggested that the information gained from comparisons made within small spatial scales may be particularly valuable to understanding the ecological variables driving primate behavioral patterns and population dynamics. Comparative investigations on

interbreeding populations living in proximate habitats are likely to limit methodological

variations between study sites, decrease the potential variations that might be caused by

phylogenetic differences between focal groups or unmeasured ecological variables, both

of which are more likely to occur at larger spatial scales (Chapman & Chapman 1999;

Chapman & Rothman 2009). When available, altitudinal gradients offer ideal conditions

to study such interspecific variations, to date, however few studies have done so.

Along an increasing altitudinal gradient, environmental conditions often shift

dramatically over short distances in relation to consistent global geophysical variables

(e.g. declines in available land area, total atmospheric pressure, temperature, and

increases in total solar radiation, and the fraction made up from UV-B radiation), as well

as local/regional climatic variables (e.g. precipitation, moisture, wind velocity, and

seasonality) (Körner 2007). As a result, species richness, diversity and productivity

decline along an increasing altitudinal gradient (Rahbek 1995; Luo et al. 2004), leading

to dramatic differences in the habitat structure and food available to organisms living

within different altitudes (Hodkinson 2005). In general, lowland habitats are

characterized by increased availability and diversity of fruit foods, whereas montane

forests have decreased fruit availability, but an increase in fiberous foods (eg. Tree ferns

and terrestrial herbaceous vegetation) (Goldsmith 2003; Ganas & Robbins 2005).

Information concerning the altitudinal variations in diet and feeding ecology of

primates is surprisingly limited. Much of what is available comes from interspecific

studies or comparisons of independent investigations of conspecific populations

(e.g.(Caldecott 1980; Goldsmith 2003; Rogers et al. 2004; Kim et al. 2011; Tsuji et al.

2013), however several interdemic studies have been performed (Byrne & Whiten 1993;

Hanya et al. 2003; Ganas & Robbins 2004; Grueter et al. 2009). These studies indicate

that primates within high elevation sites often have lower dietary diversity, and in

response to low availability or spatial distribution of preferred foods, switch to diets

consisting primarily of locally abundant food sources, such as leaves or herbaceous

vegetation. These foods, commonly referred to as “fallback foods”, are often of lower

quality than preferred foods, yet because they enable individuals to survive under sub-

optimal resource conditions, some researchers have argued that they may shape the

ecology, behavior, morphology, and divergence of species (Marshall & Wrangham 2007;

Constantino & Wright 2009; Lambert 2009; Marshall et al. 2009).

Bioko Island drills (Mandrillus leucophaeus poensis) have repeatedly been

designated over the past three decades as a relatively understudied species, and one of the

African primates most in need of conservation action (Hoshino 1985; Norris 1988;

Schaaf et al. 1990; Gonzalez-Kirchner & de la Maza 1996; Wild et al. 2005; Morgan et

al. 2013). Far fewer studies have been published on the ecology and behavior of the drill

(Gartlan, 1970; Astaras et al., 2008; Astaras, 2009) than their congeneric, the mandrill

(Mandrillus sphinx) (Abernethy et al., 2002; Hoshino, 1985; Hoshino et al., 1984; Kudo,

1987; Kudo and Mitani, 1985; Lahm, 1986; Norris, 1988; Rogers et al., 1996; Sabater Pi,

1972; Telfer et al., 2003). Information on the insular drill subspecies (M. l. poensis) on

Bioko Island, Equatorial Guinea, is even more limited. With the exception of an article

describing field notes on drills collected during a wider primate research program on the

island (Gonzalez-Kirchner & de la Maza, 1996), ecological information on Bioko drills is

only embedded within more general discussions of Bioko Island primates (Butynski and

Koster, 1994; Gonzalez-Kirchner, 1994; Mate and Colell, 1995; Hearn and Morra, 2001;

Hearn et al., 2006), all of which are mostly anecdotal in nature.

Because of the taxonomic affinity and morphological similarity of drills (M.

leucophaeus) and mandrills (M. sphinx), they have long been presumed to share similar

ecological and behavioral characteristics (Caldecott 1996; Grubb 1973), leading to broad

generalizations across the genus. However, drills are known to use a wider variety of

habitat types within a greater elevation range than mandrills. Mandrills are found almost

exclusively within closed canopy, lowland forests below 900m asl, and avoid areas

containing dense herbaceous groundcover (Kingdon et al., 2003; Lahm, 1986; Wood,

2007). Although drills are also commonly found within lowland forests, on Bioko Island

their range extends into montane forest habitat, at least 1000m asl, and hunters have

reported them up to 1500m asl (Butynski & Koster 1994; Gonzalez-Kirchner & de la

Maza 1996)(Gonzalez-Kirchner, 1994). On the mainland, drills have been reported

through the montane forest habitat, up to the montane grasslands of Mount Kupe,

Cameroon at 2000m asl (Wild, Morgan, and Dixson, 2005). Unlike what was reported of

mandrills, drills are regularly encountered in areas with dense understories of herbaceous

vegetation and grassy slopes, within both their insular and mainland ranges (Sanderson,

1940; Schaaf, Butynski, and Hearn, 1990; Wild, Morgan, Dixson, 2005).

In the first dietary analysis of drills, Astaras (2009) did not find any significant

differences between the dietary compositions of mainland drills (M. l. leucophaeus) in

Korup National Park, Cameroon, and mandrills in Campo Reserve, Cameroon, providing

some confirmation to the assumptions of their ecological similarity. Both species were

found to have primarily frugivorous diets, with fruit composing over 80% of their mean

fecal sample weight throughout the year. The consumption of the remaining components

was also relatively similar between species, although there were some minor variations

between the proportions of fibrous foods, such as leaves, shoots, and piths (drill 11% vs

mandrill 6%), animal remains (6% vs 7%), and mushrooms (2% vs 1%) (Astaras 2009).

Seasonal shifts in fruit consumption were found to correspond to fruit availability in both

species. Periods of fruit scarcity were marked primarily by increased seed depredation,

and to a lesser degree, an increased proportion of fibrous foods.

Based on comparisons of the first dietary analysis of drills by Astaras (2009), a

study that used comparable methodologies on mandrills by Hoshino (1985), and to a

lesser degree, several anecdotal comments or ad hoc accounts in the literature (Struhsaker

1969; Schaaf et al. 1990; Gonzalez-Kirchner & de la Maza 1996), three general features

of the diet and feeding ecology across the Mandrillus genus have emerged, including: 1)

broad omnivorous diets: each consuming parts from over 100 plant species and numerous

invertebrate taxa, 2) strong preference for fruit: the monthly mean proportion of fruit in

the feces remains above 60% throughout the year, reaching a maximum of more than

95% in times of high fruit availability, comprising 82.2% of drill (Astaras, 2009) and

84.2% of mandrill (Hoshino, 1985) dry fecal sample weight throughout the year, 3)

seasonal shifts in dietary composition correlated to fluctuations in fruit availability: in

times of low fruit availability, drills and mandrills increase their consumption of fallback

foods, including crushed seeds, herbaceous fiber, mushrooms, and leaves.

In this chapter I compare the diet of Bioko Island drills between montane (900-

1200 m asl) and lowland (0-300 m asl) forest sites within a small spatial scale (distance

between sites <17 km), to determine the impact of altitudinal variations in food

availability on their feeding ecology and foraging strategies. I predicted that drills would be primarily frugivorous, but that an increased consumption of fibrous vegetation and seed depredation would occur within the montane forests, in response to altitudinal

changes in fruit abundance. Further, I compare these findings with those of mainland

drills (Astaras 2009) and mandrills (M. sphinx) (Hoshino 1985), and discuss the

implications of altitudinal dietary variations on our understanding of the ecology,

behavior, and conservation of these primates.

Methods

Study Site

This study was conducted within the Gran Caldera and Southern Highlands

Reserve (GCSHR) (3.42 to 3.21 and 8.414 to 8.742 decimal degrees), one of the two nationally recognized protected areas on the island (Chapter 1: Figure 1.4). The GCSHR comprises the southern third of the island (510 km2) and is characterized by steep

altitudinal gradients, resulting from the two volcanic peaks in this region of the island,

Gran Caldera and Pico Biao, both greater than 2000m asl and within 15 km of the

coastline. The GCSHR remains one of the least disturbed and most remote areas of the

island, and the only remaining location where each of the island’s seven native monkey

species occur.

Primary feeding observations, foraging remains, and fecal samples were collected

from three locations within the GCSHR (Chapter 1: Figure 1.4). The Montane forest site

“Caldera” (900-1200m asl), within the crater of the Gran Caldera de Luba volcanic peak,

is the only remaining Montane forest habitat on the island without heavy hunting activity,

resulting from its distance from the closest road and rough terrain. Two lower elevation

sites (0-300m asl), were sampled within the western and eastern coastal Lowland forests.

“Moraka Playa”, was to the west, near the mouth of Rio Ole, and 12 km from the closest

village (Ureca). Both the Caldera and Moraka Playa are used for forest research activities

throughout the year and each contain a network of over 16 km of pre-existing trails and several rivers, all of which were used during this study. The second Lowland site,

“Moaba Playa” was to the east of the mouth of the Moaba River. This location lacks the established trail network available at the other sites, therefore surveys were performed along a 5 km trail paralleling the coastline between Punta Dolores and Punta Santiago, as well as several rivers in the area.

Food Availability Estimates

To provide a rapid estimate of the relative availability of fruiting trees between the montane and lowland forest habitats, belt transect surveys 8000 m long and 20 m wide were performed in 2011 (Sutherland 2006). During the belt surveys each individual fruiting tree within 10 m of each side of the trail, known or assumed to be consumed by drills, was counted to provide an estimate of the number of fruiting trees for an area of 16 km2 at each site.

To provide an additional estimate of the relative availability of fruit, as well as the

availability of terrestrial herbaceous vegetation, a modified version of the point quarter

quadrant method was used (Dunbar, 1983). At 100 m intervals along 6 km of the survey

transects, points were established, around which four quadrants were designated using the

N-S and W-E lines of a compass. The dominant and secondary ground cover at each

point was estimated in a 1 x 1 m quadrat, 2–5 m off the trail in a randomly chosen

direction perpendicular to the trail (Beymer and Klopatek, 1992). Groundcover type was

recorded as dirt, rock, leaf litter, fern, Aframomum sp., grass, saplings, shrub, or other

terrestrial herbaceous vegetation. Aframomum sp. was selected as it was identified from

primary observations and feeding remains collected during the exploratory field work for

this study as a potentially important food resource for drills. Within each of the four

quadrants, the tree closest to the point with a diameter at breast height (DBH) greater than

20 cm was located and the presence of fruit on the tree (at any stage) was recorded.

No survey or assessment was performed at Moaba Playa as there were no

established census trails at this location and much of the fecal sample patrol route is

either bordered on by open beaches or along rivers. However, as Moraka Playa and

Moaba Playa are within 16 km of each other and both within the lowland forests, the

overall availability of fruiting trees was assumed to be similar.

Fecal Sample Collection and Analysis

Fecal samples were collected opportunistically from wild, free-ranging, drills

within the GCSHR over a total of 3962 km of patrols during the height of the dry season

(January through March) of 2010-2012 (199 field days). Patrols and surveys were

performed using an existing trail network, rivers, beaches, and by tracking groups off-

trail using vocalizations and foraging signs. As drill groups are actively targeted

throughout the island for the commercial bushmeat hunting trade, habituation was not

attempted and as a result samples were collected from un-identified individuals.

Whole, fresh, fecal samples (n = 234) were collected, weighed, and 2g

subsamples were collected and stored for future parasite and molecular analysis. The

bolus was gently broken apart and washed with water in a 1 mm2 sieve. The water and

fecal material passing through the sieve was collected and filtered through a sheet of

cloth. This process was repeated until the wastewater was clear. The remains, both >and

< 1mm2, were wrapped separately in aluminum foil and dried in a plant drier using a paraffin stove until a steady weight was achieved. The samples, still wrapped in their foil boats, were placed in individual Ziploc bags, stored in a pelican case with silica gel desiccants, and transported back to the laboratory at Drexel University for analysis. The dry fecal remains were then sieved through 25 mm2, 4 mm2, and 1 mm2 mesh sizes,

which simplified the process of identifying and categorizing each individual particle

greater than 1 mm2. These remains were separated into the following dietary categories:

Non-fruit Fiber (including shoots, roots, and piths), Fruit Fiber (including skin and flesh),

Seed, Leaf, Animal (including vertebrates and invertebrates), Mushroom (sporocarps),

and Others (including wood, twigs, and soil). The weight of each category was recorded

(Mettler, model AE240, +0.001g), however seeds >5mm diameter were not included in

this measurement, to avoid overestimating Seed contribution to their diet (Hoshino 1985).

To estimate the relative volume of the remains of each category, the remains were spread

on 1cm2 grid paper at a consistent height and density, and the number of cells covered (+

0.5%) by each category counted (Astaras, 2009).

Seed and Animal remains >1mm2 were identified to the lowest possible taxonomic

group whenever possible. This was not done on the other food categories as their remains

typically lacked taxonomically identifiable characteristics. Unidentified remains were

assigned a distinct code, and all distinctly similar unknown remains were grouped into

one morphotype and included in the total count of food items eaten (n) in each site

(Astaras, 2009).

Data Analysis

The dietary diversity (H’) was calculated by site for both Seed and Animal

remains within the feces using the Shannon-Weiner diversity index: ′ ∑ ∗ .

The frequency of occurrence for each food item (Oi) was used to calculate its relative

frequency (Pi) in the diet at each site through: /∑. Dietary evenness (J’) was

calculated for the Shannon-Weiner measure, to provide a relative measure from 0 to 1

based on the n of each site, using: ′ ′/ln (Cords 1986). To test for differences in

the presence and absence of Seed and Animal remains between the three sites, individual

Pearson’s Chi-squared analyses (two-tailed) were used.

After Shapiro-Wilk test indicated that non-parametric analyses were appropriate

for statistical comparisons, separate Kruskal-Wallis tests were performed to compare the

weight and volume of each food category by site. Subsequent multiple comparisons were

made using kruskalmc from R package pgirmess (Giraudoux 20012). Pairwise

comparisons of the dietary overlap between the three study sites were made for the

weight and volume values of the fecal remains separately, using Czekanowski’s

proportional similarity index (PS) (Schoener 1968): 1 0.5 ∑| |.

Where Pij and Pik are the frequencies of the food category i, in the diets at sites j and k,

for the total number of food categories, n. Dietary overlap (PS) provides a simple

measure from 0 (the absence of overlap) to 1 (complete dietary overlap).

To determine if the consumption of either fruit or fiber was proportional to its

relative availability at a given site, a dietary preference index (D) was calculated.

Relative consumption (i) was estimated using the average proportion of the fecal

sample mass at the Caldera (c) and Moraka Playa (m) composed by fruit and non- fruit

fiber remains. The relative availability () of these food items in the habitat were based on the frequency that fruiting trees and terrestrial herbaceous vegetation (including

Aframomum sp.) were present at each habitat assessment point along the Santo Antonio and Badja Trails. The metrics used to assess the availability of each food type are non-

comparable, however they remain proportionally relative between sites. Thus, the analysis was performed for fruit and fiber separately.

The index was calculated as: /⁄ ̅/̅, where D = 1 indicates a lack of preference for that food category (i.e. consumption is proportional to its relative availability within both habitats). Values less than one indicate a higher preference of that food type in the Caldera, and those greater than one a preference at Moraka Playa.

To generate confidence intervals for D and determine if the results were significantly different than a null model, in which dietary preference is independent of the collection site, a bootstrap approach was used. For each variable, and ̅, new sample populations that were equal in size to the original were generated for each site by resampling with replacement from the observed data. D was calculated for each new population, and repeated for 10,000 iterations. Under the null model, the bootstrap data for each site was resampled randomly from both the Caldera and Moraka Playa, whereas in the alternative model bootstrap data was resampled from within each site. The 95%

confidence intervals of the mean D values were calculated and compared between the two models to determine significance of the observed values.

Principal Component Analyses (PCA) were performed to study the patterns of dietary variation at each site. The weight and volume values of each dietary component

(non-fruit fiber, fruit fiber, seed, animal, leaf, mushroom, and other) were log10

transformed, and analysis was carried out on the correlation matrices. The scores of the

first three principal components were used in Multivariate Analysis of Variance

(MANOVA) to compare dietary variables between sites. Subsequent site comparisons

were made through pairwise one-way ANOVAs.

All analyses were performed using R statistical software v 2.15.0 (R Core

Development Team 2012). Significance tests were two-tailed with the significance level

set at 0.05.

Ethical Note

This research complied with the all legal requirements of the government of

Equatorial Guinea and the safety and ethical protocols of the American Society of

Primatologist’s Principles for the Ethical Treatment of Nonhuman Primates. The non-

human primate fecal samples used in this study were handled, stored, and transported

under the guidelines of the US Fish and Wildlife Services and US Center for Disease

Control.

Results

Food Availability Estimates

Estimates of fruit availability via belt transect and point quarter methods show a distinct

difference in fruit available between montane and lowland forest types. A total of 185

fruiting trees (11.56 trees/km2) were counted along the belt transect at Moraka Playa,

only two (0.13 trees/km2) were counted within the Caldera. The mean presence of

fruiting trees within each quarter of the point quarter assessment was significantly higher

at Moraka Playa (Fisher’s Exact Test, p < 0.001), where 44.3% of points had at least one

of four trees with fruit present, compared to a single fruiting tree that was encountered

within the Caldera (0.54% of all the trees assessed in the montane forest).

Both the dominant and secondary groundcover types were found to differ

significantly between sites (Fisher’s Exact Test, each at p < 0.001). Leaf litter made up

the majority of the dominant groundcover within the 6 km interval at both locations.

However, the points at Moraka Playa had higher rock and other herbaceous plants (e.g.

Paracostus englerianus and Dracaena phrynioides), and the Caldera had more

Aframomum sp. and grasses. The secondary groundcover types with the highest

frequency in the Caldera were Aframomum sp., (41%) and fern (28.3%), whereas leaf

litter (30%), rock (28%), and saplings (28%) were highest in the lowland forest was.

Qualitative Dietary Analysis

A total of 234 fecal samples were collected during the study period, 129 from the

Caldera (mean fresh weight = 61.7 g, SD= +40.5), 81 from Moraka Playa (mean fresh

weight = 28.8g, SD= +25.5), and 24 from Moaba Playa (mean fresh weight = 37.5g, SD=

+23.8) (Table 2.1). Mean fresh fecal weight was significantly higher within the Caldera

compared to the Lowland sites (Kruskal-Wallis, H=39.86, p<0.001).

I identified plants from 50 distinct taxonomic groups (39 species and 11

morphotypes), from at least 34 genera and 23 families, as drill foods from the fecal

sample analysis and field observations (Table 2.2). The list of food items eaten at the

lowland forest sites, Moraka Playa and Moaba Playa, were dominated by the fruits of 17

species of trees (77.3% of the identified vegetative species eaten), a shrub (Leea

guineensis)(4.6%), and a vine (Momordica foetida)(4.6%). Only three species (13.6%) of herbaceous vegetation (from the family Costaceae), of which they ate the stem pith and roots, were consumed by drills in the lowland forests. In stark contrast to the lowland sites, fruiting trees only constituted 6 of the 20 (30%) identified food species consumed by drills in the Caldera. Within this montane forest location, drills consumed food items from a greater variety of plant types and parts, including: the stems, roots, flowers and fruits of seven herbs (35%), the fruits and leave petioles of 2 shrubs (10%), the stems, leaves, and grains of two grasses (10%), the stems of a sedge (5%), shoots of one fern

(5%), and the viscous pith of the tree fern, Cyathea manniana (5%).

The frequency of seed presence (Oi) within the fecal remains found in the Caldera was significantly lower (0.6) than either Moaba Playa (0.85) or Moraka Playa (0.82)

(Pearson’s Chi-squared, X2=10.07, df=2, p=0.0065)(Table 2.3). The diversity (H’) of

seeds found in the fecal samples was highest at Moraka Playa (H’=1.73), followed by the

Caldera (H’=1.28), and then Moaba Playa (H’=1.16). However, Moaba Playa had the

highest evenness index (J’) for seed consumption (Moaba=0.84, Moraka Playa= 0.79,

Caldera=0.52). The disparity between H’ and J’ at Moaba Playa likely due to the

relatively low number of samples collected (N=20) at this site.

Through fecal analysis and field observations we identified 20 distinctly different

animal taxa eaten by drills, which included three identified species and 17 morphotypes

representing 11 taxonomic orders (Table 2.4). Within the Caldera fecal samples,

Coleopterans (Pi=34%), Hymenopterans (Pi=16%), and African giant millipedes

(Archispirostreptus gigas) (Pi=20%) were the most frequently present animal remains.

Drills in the Caldera were frequently observed breaking adult millipedes in half to suck

out the innards, a behavior only observed at this site. Land crab (Johngarthia weileri)

were identified as a particularly important food source at Moaba Playa, where (55%) of

samples contained crab shell. Surprisingly, crab remains were only found in 6% of

samples from Moraka Playa, despite there being no obvious difference in the abundance

of crabs between these two locations. Moraka Playa had the highest taxonomic richness

of animal remains (12), including the only vertebrate remains found during this study.

Skeletal material from a small rodent (Familiy: Muridae) and a frog were found within a

single fecal sample at this site.

The frequency of animal remains present in the fecal remains was significantly

higher within the Caldera (0.94) and Moaba Playa (0.85), compared to Moraka Playa

(0.6) (X2=23.44, df=2, p<0.0001)(Table 2.3). Dietary diversity of animal consumption

was highest at Moraka Playa (H’=1.73), followed by the Caldera (H’=1.28), and Moaba

Playa (H’=1.16). Dietary evenness was highest within the Caldera (J’=0.82) compared to

the lowland forest sites, which had equal evenness values (J’= 0.78).

Quantitative Dietary Analysis

Quantitative dietary analysis was performed on 150 fecal samples (Caldera N=59,

Moraka Playa N=71, Moaba Playa N=20). Significant dietary differences were found

between the study sites in the mean values and relative proportions of the food categories

(Figure 2.1). The differences in food categories between sites remained the same for the

weight and volume metrics, thus we will only further discuss the comparisons made by

weight, except where otherwise noted.

Drill fecal samples collected in the Caldera were primarily composed of non-fruit

fiber (59.4%, SE=4.29), which was significantly higher than Moraka Playa (5.7%,

SE=2.29) or Moaba Playa, where non-fruit fiber was not found in any samples (Kruskal-

Wallis, H=85.09, p<0.001). Foraging observations of drills during this study indicate

that the majority of the non-fruit fiber within the Caldera primarily originated from the

stem-pith and roots primarily from four species from the genus Aframomum (Table 2.2).

Feeding on these herbs involved biting off a section of the stem, which the drills would

then split apart with their hands to access and eat the center-most pith. Drills of all age-

classes, except infants, and both sexes have been observed foraging within large stands of

Aframomum, the remains of which were found in 89.8% of fecal samples collected in the

Caldera.

Fruit weight was significantly higher in fecal samples collected at Moraka Playa

(88%, SE=3.08) and Moaba Playa (91.4%, SE=3.59), compared to the Caldera (21.0%,

SE=3.92) (Kruskal-Wallis, H=75.19, p<0.001). In comparing fruit consumption between

the lowland sites, fruit fiber weight was significantly higher at Moaba Playa (59.7%,

SE=6.3 vs 31.7%, SE=4.36), whereas seed weight was significantly higher at Moraka

Playa (56.3%, SE=4.82 vs 31.7%, SE=6.68) (Kruskal-Wallis, Fruit Fiber: H=10.07,

p=0.005; Seed: H=5.34, p=0.02).

Animal remains made up a significantly higher proportion of the diets at Moaba

Playa (3.1%, SE=1.01) compared to Caldera (2.0%, SE=0.54) or Moraka Playa (2.2%,

SE=1.36) (Kruskal-Wallis, H=30.0, p<0.001). Leaves made up significantly more of the

diet of drills within the Caldera (7.6%, SE=2.05) compared to Moaba Playa (4.4%,

SE=3.31) or Moraka Playa (0.8%, SE=0.25) (Kruskal-Wallis, H=47.5, p<0.001).

Through primary observations and secondary feeding encounters following a foraging

bout, drills in the Caldera were found to consume the leaves of nine species of plants,

many times in conjunction or subsequent to eating another part of the plant, such as the

stem, fruit, or flower. A significantly higher proportion of fecal samples of drills in the

Caldera contained mushroom (7.0%, SE=1.94), compared to Moraka Playa (0.5%,

SE=0.18) or Moaba Playa (0%)( Kruskal-Wallis, H=50.3, p<0.001). The proportion of

the category Other, which was primarily composed of wood, was significantly lower at

Moaba Playa (1.06%, SE=0.99) as compared to the Caldera (3.1%, SE=1.0) and Moraka

Playa (2.8%, SE=1.1) (H=8.1, p=0.02).

Principal components analysis (PCA) of the weight values of the dietary

categories generates three components that explain 72% of the total variance (Table 2.6).

The first component contrasts diets high in Fruit Fiber, and to a lesser extent Seed, with

those in which Non-fruit Fiber, Leaf, and Mushroom are the most important categories.

The second component contrasted diets with high proportions of Seed and Other with those high in Fruit Fiber and Animal. The third component contrasts diets high in Fruit

Fiber and Animal with those where only Seed contribution is of high importance. When

comparing the scores of the first two principal components for each individual fecal

samples, two distinct dietary clusters emerge; the first in the Caldera, characterized by

Non-fruit Fiber, Leaf, and Mushroom, and the second within the lowland sites,

characterized by Fruit Fiber, Seed, and Other (Figure 2.2).

The scores of the first three components are significantly different when

comparing the main effect of site (MANOVA, F2,147=32.8, p<0.001). Subsequent

univariate F-tests for each of the principal components indicate that much of this difference is in the first and third components; however the second was still significant

(PCA1:F2,147=102.7, p<0.001; PCA2:F2,147=3.3, p=0.04, PCA3:F2,147=16.1, p<0.001). Site

comparisons of the scores were made through independent pairwise ANOVAs. PCA1

scores are significantly different between the Caldera and both lowland sites (Moraka:

F1,128=163.2, p<0.001; Moaba: F1,77=89.3, p<0.001). PCA2 scores are marginally

significant between Caldera and Moraka Playa (F1,128=5.6, p=0.02). Finally, PCA3

scores of Moaba Playa were significantly different than either of the other sites (Caldera:

F1,77=20.8, p<0.001; Moraka: F1,89=30.4, p<0.001).

Dietary overlap, as indicated by Czekanowski’s proportional similarity index

(PS), was high between the lowland sites (0.92). Caldera drill diets overlapped little with

those of Moraka Playa (0.33) or Moaba Playa (0.29) (Figure 2.3). Dietary preference

index (D) for fruit (0.15) and fiber (0.33) consumption was significantly higher in the

samples collected in Caldera, compared to those in Moraka Playa. The 95% confidence

intervals of the mean D values of the alternative bootstrapped datasets fell outside those

of the null models, indicating that these observed preferences were significant (Table

2.5).

Discussion

Bioko Island drills exhibit two distinct dietary patterns within the Gran Caldera and Southern Highlands Scientific Reserve. Within the lowland forest sites they utilized a similar feeding strategy to the mainland Mandrillus; although they consumed a wide variety of food types, fruit comprised 88% of their fecal remains, and 77% of the identified vegetative food consumed were from fruiting tree species (Table 2.7). In contrast, drills in the montane forest were primarily folivorous, as 67% of their fecal

remains were composed of non-fruit fiber and leaf, and 60% of the plant foods were from

shrub, herb, and grass species. These results also indicate that drills compensate for this highly folivorous diet by increasing their consumption of insects, mushrooms, and leaves.

We attribute these differences found on Bioko Island to the dramatic variations in the habitat structure between montane and lowland forests, especially in regard to the inverse relationship between the relative availability of fruiting trees and herbaceous groundcover between the two forest types.

In general, the response of drills within the montane forests of Bioko Island to low fruit availability was substantially different than that reported of the mainland

Mandrillus during fruit lean period (Hoshino 1985; Astaras 2009). In light of the relatively low availability of fruit in the montane forests of Bioko Island, one would expect high seed consumption, and although whole seeds were prevalent in the fecal samples at all three sites (Caldera = 60%; Moraka Playa = 82%; Moaba Playa = 85%), crushed seeds were never encountered in the 150 fecal samples analyzed during this study.

Seeds are typically thought to represent a particularly important food resource for

mainland drills and mandrills. The presence of crushed seeds in the fecal samples of

mainland drills was found in an average 94.9% of samples throughout the year (monthly

range 79-100%), and made up 25% of the annual mean dry weight of their feces (Astaras

2009; Astaras & Waltert 2010). Seeds are more resistant to decomposition than other fruit

tissues, and so they persist in the leaf litter beyond the fruiting period (Fleagle &

McGraw 2002), explaining the negative correlation between seed depredation by drills

and mandrills and fruit availability throughout the year (Jouventin 1975; Rogers et al.

1996; Tutin et al. 1997; Astaras & Waltert 2010). As members of the Cercocebus-

Mandrillus clade of the Papionins, drills are characterized by several unique adaptations

for terrestrial foraging in the leaf litter and masticating hard seeds and nuts (Fleagle &

McGraw 1999). An increased ability to exploit seeds may provide a critical means of

surviving during periods of fruit scarcity, particularly where interspecific frugivore

densities, and the resulting competition, are high. Thus, the unique adaptations of the

Cercocebus-Mandrillus clade likely reflect an evolutionary response to seeds as a

fallback food (Marshall & Wrangham 2007).

Given this adaptation to seed consumption and the reported use of seeds as

fallback foods, the switch to a diet primarily composed of the pith of terrestrial

herbaceous vegetation by Bioko Island drills in the Caldera was unexpected. Seasonal

consumption of the stem pith of Aframomum sp. and other herbs has been reported for

both drills (Astaras 2009) and mandrills (Hoshino 1985; Lahm 1986; Norris 1988),

however it has never been found to constitute more than 25% of their total diet in any

month.

The relative proportion of animal content in the fecal samples was highest at

Moaba Playa, however the majority (61%) of the remains were from the exoskeleton of

crabs, which have relatively high weight and volume compared to the remains of the

other invertebrate foods. There was no evidence from fecal sample analysis or field

observations that drills at Moraka Playa consumed crabs, despite both sites being

adjacent to coastal habitat and no obvious difference in land crab abundance between the

two sites. These results are likely to disproportionately over-represent the relative amount of animal tissues consumed at Moaba Playa. For example, drills in the Caldera are regularly seen feeding on millipedes and orthopterans, and observed stripping the leaves from the shrub, Anthocleista vogelii, to search the petioles for sawfly larvae (Order:

Hymenoptera; Suborder: Symphyta). These small (~1.5 cm) larvae have soft bodies, which would likely leave little to no remains in the feces of a drill; thereby underrepresenting the Animal dietary component of this individual. Drills at Moaba Playa were rarely observed searching rotten logs for invertebrates, a common behavior at

Moraka Playa, the Caldera, and at Korup National Park (Astaras 2009), likely because much of the animal diet at Moaba Playa was composed of land crabs.

Mushrooms constituted a significantly higher proportion of the samples in the

Caldera, and was 5-6% higher than in the diets of drills or mandrills on the mainland

(Hoshino 1985; Astaras 2009). The consumption of mushrooms is relatively rare in wild primate diets, as it has been recorded in only 23 species (Hilário & Ferrari 2011) and constitutes less than 5% of the feeding time of the majority of these consumers(Hanson et al. 2003). Fungi are considered a low quality food source for most primates, as their digestion often requires the consumer to have specialized fermentation capabilities to access nitrogen and carbohydrate stores (Hanson et al. 2006). As a result, where

mycophagy is recorded, it is often primarily associated with low fruit availability, which is consistent with this study and previous reports of the mainland Mandrillus spp.

(Astaras 2009).

Although the relatively high proportion of non-fruit fiber, insect, mushroom, and

leaf in the fecal samples of the drills in the Caldera may be explained by differences in

the habitat, the complete absence of seeds at any of the sites on Bioko cannot be. This is

also true of the difference in crab consumption between Moraka Playa and Moaba Playa,

as drills at both sites live in forests bordering the beach, both with large numbers of crabs

(pers. obs.). Even if differences in the availability of these foods did exist, it is unlikely

that they, alone, would explain the complete absence of crab consumption at only one

location. Throughout the primate literature there are numerous records of dietary

differences between groups and populations that cannot readily be explained by

environmental variations between local habitats, such as the relative abundance or

distribution of prey items or competition (McGrew 1983, 1998). Several alternative

hypotheses have been proposed to account for the differences; most common are the

relative profitability of the food items (i.e. the food choice based on the nutritional

benefits and energetic costs of foraging) or the selection of foods based of certain physical characteristics or non-nutritional secondary compounds for medicinal purposes

(treatment of parasitic infections) (Chapman & Fedigan 1990; Huffman 1997). In fewer cases, such variations have been best explained by local cultural traditions, where new or modified behaviors are transmitted to, and consistently used by, other individuals, which then persist throughout multiple generations (McGrew 1998).

Much of the evidence for the existence of cultural traditions in non-human

primates comes from the great apes, while the literature has largely ignored cultural

traditions in non-Hominid species (McGrew 1998). Studies of captive chimpanzees (Pan

troglodytes) have demonstrated their ability to transmit and sustain learned behaviors

between and within groups (Whiten et al. 2007), however such hypotheses have been

difficult to confirm in the wild because of the effects of unknown, or untested,

environmental variables, which can never be completely excluded (McGrew 1983).

One of the most compelling examples of cultural dietary differences in wild

primates is in the variable consumption of insects by western lowland gorillas (Gorilla

gorilla gorilla) at five study sites in Central Africa (Deblauwe & Dupain 2003). Ants and

termites were commonly eaten by groups at two of the sites (Ndoki, Congo, and Ntonga,

Cameroon); those at another two sites (Belinga, Gabon, and Dzanga-Sangha, CAR)

consumed termites, but not ants, while the opposite was true for groups at Lopé, Gabon.

Deblauwe and Dupain (2003) suggest that these dietary variations are most likely a result

of local cultural traditions, as the availability of neither ants nor termites differed appreciably between any of these sites. This finding is consistent with the differences in seed consumption between drill populations, and interdemic differences in crab consumption, thus cultural differences may provide the best explain for these differences.

However, additional research is needed to assess the relative availability of these foods between each location, as well as any potential nutritional, or foraging benefits that might explain these differences.

Diet and Conservation

Primates live in an increasingly changing world. The primary threats to primate

species worldwide, climate change, habitat destruction, human encroachment and

hunting, all have the potential to modify the remaining habitats available to wild

populations (Cowlishaw & Dunbar 2000). How primates will respond to these threats is

largely dependent on the plasticity of their unique ecological traits and behavioral

characteristics (Chapman & Onderdonk 1998), therefore a focus should be placed on

research efforts to better understand these dynamics at multiple scales.

Not only is this information important for species conservation, it is also critically

important for ecosystem management. Primates have been found to make up a large

portion of the mammalian frugivore biomass in numerous tropical forest sites (Eisenberg

& Jr 1973; Prins & Reitsma 1989; Oates et al. 1990; White 1994; Chapman & Onderdonk

1998), and, as such, can be vital to the maintenance of their forest ecosystems. This is

particularly true of the role primates play in seed distribution (Chapman 1995; Dew and

Wright, 1998; Chapman and Onderdonk, 1998; Poulsen et al., 2002). However, we found

the seed handling and depredation between mainland and insular drills to differ

significantly, implying that the extirpation of one drill population might have

dramatically different ecological consequences than the loss of another.

Conclusions

Several studies have recently stressed the importance of dramatic ecological

variations that can exist between species and populations within small spatial scales.

(Butynski 1990; Chapman & Fedigan 1990; Chapman & Chapman 2000b; Chapman et al. 2002). Within a single population of drills separated by less than 20 km, we found that natural variations in food availability between montane and lowland forests significantly

changed the diet and feeding strategies of drills. Our results also show that some of the

long-standing assumptions concerning the ecological and behavioral similarities between

drills and mandrills were not ubiquitously applicable. The dietary differences reported

here between Bioko drills in montane and lowland forest habitats is similar to what has

been reported for other omnivorous primates, including gorillas (Gorilla spp.)(Doran &

McNeilage 1998; Ganas & Robbins 2005) and Japanese Macaques (Macaca

fuscata)(Hanya et al. 2003), indicating that studies performed on populations living along

elevation gradients may be critical to a more complete understanding the factors driving

intraspecific ecological and behavioral variation.

7 250 Caldera 6 200 Moaba Playa

5 ) 2 Moraka Playa 150

(g) 4 (cm

3 100 weight

2 volume 50 1

0 0 100 100

75 75 total) total)

of of

50 50 (% (%

25 25 weight volume 0 0

Figure 2.1: Mean weight (left) and volume (right) of the dietary categories by their absolute values (top) and proportion within the total diet (bottom).

Figure 2.2: Bivariate plot of the first two principal components scores for each location.

Figure 2.3: Pairwise comparison of the dietary overlap between the sites, as indicated by Czekanowski’s proportional similarity index (PS).

Table 2.1: Fecal sample collection effort and results for Bioko Island drills at the three study sites. Mean Weight Location Effort (km) Survey Days Fecal Samples Samples/km Samples/Day (Range) Gran Caldera 1407 135 129 0.09 0.96 61.7g (1.9-197.7) Moraka Playa 1316 45 81 0.06 1.80 28.8g (1.1-119.1) Moaba Playa N/A* 28 27 N/A* 0.96 37.5 g (2.9-80.1) *Samples were collected primarily following observations made in blinds, patrols surveys were entirely on unmarked trails, rivers, or coastlines, thus no distance estimates are given.

Table 2.2: List of vegetative food items consumed by Bioko Island drills in this study. Family Species Form1 Parts 2 Site3 Anacardiaceae Spondias sp. T Fr Moraka Annonaceae Annona muricata T Fr Moaba Annonaceae Monodora myristica T Fr Moraka Annonaceae Xylopia aethiopica T Fr Lowland Apocynaceae Hunteria umbellata T Fr Moraka Apocynaceae Landolphia sp T Fr Moraka Araceae Anchomanes difformis H SP Caldera Arecaceae Cocos nucifera T Fr Lowland Arecaceae Elaeis guineensis T Fr Lowland Asparagaceae Dracaena phrynioides H R Caldera Burseraceae Dacryodes macrophylla T Fr Caldera/Moaba Costaceae Costus afer H Sp Moaba Costaceae Costus dinklagei H Sp Moaba Costaceae Paracostus englerianus H R Moraka Cucurbitaceae Momordica foetida V Fr Moraka Cyatheaceae Cyathea manniana TF P Caldera Cyperaceae Cyperus sp. SE Sp Caldera Dryopteridaceae Didymochlaena truncatula F L Caldera Flacourtiaceae Oncoba glauca T Fr Moaba Gentianaceae Anthocleista vogelii T Lp/Fr Caldera Graminae Puelia ciliata G St/Se Caldera Graminae Setaria megaphylla G St/R Caldera Humiriaceae Sacoglottis gabonensis T Fr Moaba Icacinaceae Lavigera macrocarpa T Fr Moraka Moraceae Ficus conraui T Fr Caldera Moraceae Ficus lutea T Fr Caldera Moraceae Ficus natalensis T Fr Caldera Moraceae Ficus sur T Fr Caldera Moraceae Ficus vogeliana T Fr Lowland Myristicaceae Coelocaryon preussii T Fr Moraka Phyllanthaceae Protomegabaria stapfiana T Fr Caldera/Moraka Phyllanthaceae Uapaca guineensis T Fr Moaba Sapotaceae Chrysophyllum africanum T Fr Lowland Vitaceae Leea guineensis SH L/Fr/Fl Caldera/Moraka Zingiberaceae Aframomum limbatum H Sp/Fr/Fl Caldera Zingiberaceae Aframomum mala H Sp/Fr/Fl Caldera Zingiberaceae Aframomum melegueta H Sp/Fr/Fl Caldera Zingiberaceae Aframomum sp. H Sp/Fr/Fl Caldera Zingiberaceae Renealmia mannii H Sp Caldera Morphotype 1 Unknown UN Se Caldera

Table 2.2: continued Family Species Form1 Parts 2 Site3 Morphotype 2 Unknown UN Se Caldera Morphotype 3 Unknown UN Se Caldera Morphotype 4 Unknown UN Se Caldera Morphotype 5 Unknown UN Se Moraka Morphotype 6 Unknown UN Se Moraka Morphotype 7 Unknown UN Se Caldera Morphotype 8 Unknown UN Se Caldera Morphotype 9 Unknown UN Se Caldera Morphotype 10 Unknown UN Se Caldera Morphotype 11 Unknown UN Se Moraka 1) F: Fern; G: Grass; H: Herb; SE: Sedge; SH: Shrub; T: Tree; TF: Tree fern; UN: Unknown; V: Vine 2) Fl: Flower; Fr: Fruit; L: Leaf; Lp: Leaf Petiole; P: Pith; R: Root; Se: Seed; Stem Pith: Sp; St: Stem 3) Lowland: Moraka Playa and Moaba Playa

Table 2.3: Comparison of the prevalence, species richness, diversity, and evenness of seeds and invertebrates within the fecal samples at each field site. Seeds Invertebrates Site Prevalence Richness Diversity Evenness Prevalence Richness Diversity Evenness (Oi) (N) (H') (J') (Oi) (N) (H') (J') Caldera 0.60 12 1.28 0.52 0.94 9 1.81 0.82 Moaba 0.85 4 1.16 0.84 0.85 4 1.09 0.78 Moraka 0.82 9 1.73 0.79 0.60 12 1.95 0.78

Table 2.4: List of animal food items consumed by Bioko Island drills in this study. Order Suborder Family Species Site Invertebrate Taxa Annelida Oligochaeta - - Caldera Coleoptera Polyphaga Elateridae - Moraka Coleoptera Polyphaga Scarabaeidae - All Sites Coleoptera Polyphaga Scarabaeidae - Caldera/Moraka Coleoptera Polyphaga Staphylinidae - Caldera/Moraka Coleoptera - - - All Sites Decapoda Brachyura Gecarcinidae 1 Moaba Decapoda Brachyura Potamonautidae 2 Caldera Decapoda Caridea Macrobrachium - Caldera Heteroptera Hemiptera - - Caldera/Moraka Hymenoptera Apocrita Formicidae - Moraka Hymenoptera Symphyta - - Caldera Hymenoptera - - - All Sites Hymenoptera - - - Moraka Isoptera - - - Caldera Lepidoptera - - - Caldera Neuroptera - - - Moraka Spirostreptida Spirostreptidea Spirostreptidae 3 Caldera

Vertebrate Taxa Anura - - - Moraka Rodentia Myomorpha Muridae - Moraka 1: Johngarthia weileri 2:Sudanonautes sp. 3: Archispirostreptus gigas

Table 2.5: Dietary preference index (D) for the consumption of fruits and terrestrial herbaceous vegetation. Values less than one indicate a higher preference within the Caldera. Confidence intervals (95%) for the null and alternative models were derived through bootstrap analysis. Food Type Mean observed D Ho mean (CI) Ha mean (CI) Fruits 0.15 1.07 (0.43-2.17) 0.16 (0-0.59) Fibrous foods 0.33 1.05 (0.54-1.9) 0.36 (0.09-0.82)

Table 2.6: Principal component scores of the dietary category masses. Food Category PC1 PC2 PC3 Non-fruit Fiber 0.510 -0.106 0.179 Fruit Fiber -0.414 0.299 -0.490 Seed -0.210 0.640 0.423 Animal 0.314 0.285 -0.689 Leaf 0.423 0.154 -0.176 Other 0.248 0.621 0.189 Mushroom 0.431 0.008 0.090 % Explained Variance 42.4 16 13.4

Table 2.7: Comparison of the methodologically comparable dietary studies on drills and mandrills, including the elevation range, fresh fecal sample weights, and mean percentages of the dietary categories for each study site. Percent dry weight, Mean, SD Fecal Mass Mandrillus Elevation Site Mean, SD Fiber species (m asl) (range) Fruit & Animal Mushroom Others Leaf 62.2 +40.5 M. l. poensis Caldera 900-1200 21.0 +30.1 67.0 +30.2 1.99 +4.2 7.0 +14.9 3.1 +8.1 (1.9-197.7)

28.8 +25.5 M. l. poensis Moraka 0-300 88.0 +26.0 6.6 +20.2 2.2 +11.5 0.5 +1.5 2.8 +9.3 (1.1-119.1)

37.5 +23.8 M. l. poensis Moaba 0-100 91.4 +16.1 4.4 +14.8 3.1 +4.5 0.0 1.1 +4.4 (2.9-80.1)

24.4 +15.7 M. l. leucophaeus Korup1 0-500 82.2 +15.3 10.7 +11.4 5.5 +6.1 1.6 +6.8 -- (1.1-93.3)

M. sphinx Campo2 50-230 -- 84.2 +13.6 5.9 +8.7 7.6 +8.4 1.0 +3.8 0.3 +0.8

1) Korup National Park, Cameroon, from Astaras (2009) 2) Campo Reserve, Cameroon, from Hoshino (1985)

CHAPTER 3: GASTROINTESTINAL PARASITIC INFECTIONS

Introduction

The potentially strong influence of parasitic infections on the maintenance of host populations is clear (May & Anderson 1978). The pathogenic effects of parasites have

been shown to reduce growth rates, impede metabolic processes, and ultimately reduce

the survival rate and fitness of their hosts (Minchella & Scott 1991; MacRae 1993;

Delahay et al. 2009). These effects may be exacerbated in the presence of additional

environmental perturbations, such as fluctuations in resource availability, temperature, or

rainfall (Pedersen & Greives 2008; Martinez & Merino 2011). In light of their potentially

strong influence on host populations, parasitic infections have increasingly been

identified as an important consideration to the conservation and management of species

(Scott 1988; Lyles, Ann M and Dobson 1993; Stoner 1996; Nunn & Altizer 2005; Mbora

& Munene 2006; Nunn 2006)/

In addition to the direct threats parasites can pose to their host populations,

preexisting or emergent parasitic infections may cause additional or secondary declines,

adding additional conservation and management concerns (Daszak 2000; Nunn 2006).

For example, populations of wild giant pandas (Ailuropoda melanoleuca) were

dramatically reduced from 1970 to 2000 due to habitat loss and degradation, food

availability shortages, and illegal poaching (IUCN 2013). Conservation programs and

policies were successful in resolving these threats, resulting in a fourfold increase in the

density of the current population within the limited available habitat remaining (Zhang et

al. 2008). Since 1980, wild panda mortalities attributed to heavy nematode infections,

particularly from Baylisascaris schroederi, have increase significantly, accounting for

half of the reported deaths from 2001-2005, which Zhang et al. (2008) attribute to a

density dependent increase in the frequency of nematode transmission.

Parasitic infections have also been shown to have significant impacts at the community level (Minchella & Scott 1991). This was effectively demonstrated by the response of the Hawaiian avifaunal community to the early 20th century introduction of

malaria (Plasmodium relictum). The emergent malarial infections resulted in the

extinction of at least nine avifaunal species and dramatic range restrictions in many of the

extant community members (Van Riper III et al. 1986). Despite the potentially high

detrimental impact of parasites to the preservation of threatened species and natural

communities, parasitic diseases are too often ignored by conservation biologists and

managers (Lyles, Ann M and Dobson 1993).

In addition to the conservation threats parasites impose on non-human host

populations, infection of wildlife populations have also been recognized as important

considerations to human health (Daszak 2000). Numerous domesticated and wild taxa

have been shown to act as reservoirs for zoonotic parasites and may ultimately promote

their transmission to human communities (Murray et al. 2000; Legesse & Erko 2004;

Gortázar et al. 2007). The zoonotic and anthropozoonotic threats are particularly high

where wildlife and human communities are in close proximity, general health standards

are low, and inadequate measures exist for the prevention of parasitic infections(Wallis &

Lee 1999). Such interactions are especially likely within the workforce of many of the

extractive industries throughout tropical forests worldwide. As people venture deeper into

the newly deforested areas, they may be introduced to pathogens to which they lack

immunity, and are thus more vulnerable to infection (Patz et al. 2000). Ultimately, as the

human population continues to expand and encroach into wildlife habitats, the risk of

zoonotic parasitic infections is likely to rise, as has been best demonstrated with such

emerging infectious diseases as Ebola virus and human immunodeficiency virus (HIV)

(Daszak 2000).

Primates have been identified as an important group for parasitic studies for

several reasons. First, the phylogenetic proximity of primates to humans make them one

of the most serious concerns for zoonotic transmission (Wolfe & Daszak 2005), which

has been well demonstrated by the impact of the aforementioned infectious viruses, Ebola

and HIV (Guenno et al. 1995; Benavides et al. 2012). Second, the often tight social

organizations and behaviors exhibited by many primate species (e.g. grooming, play,

aggression) increases the risk of parasite transmission, making them particularly

vulnerable to parasitic infections (Stoner 1996; MacIntosh et al. 2012). Third, roughly

half of all known primate species are threatened globally, primarily due to habitat loss

and bushmeat hunting, practices which increase the encroachment of humans into already

restricted rainforest habitats, as well as the likelihood of anthropozoonotic parasite

transmission to the remaining primate communities (IUCN 2013). Fourth, the relative

lack of seasonal temperature fluctuations in the tropics has been shown to result in

disproportionately high selective pressures imposed by parasites on tropical avian

species, compared to those in temperate zones (Møller 1998). Given that approximately

90% of primates live within the tropics (Mittermeier 1988), it is likely that parasites may

have had a greater role in driving the evolution of behavioral or morphological traits of

primates than other taxonomic groups less concentrated in this zone.

Although many times parasitic infections are asymptomatic in primates, hosts

may suffer from numerous manifestations, including malnutrition, tissue destruction, secondary bacterial or viral infections, and sometimes death (Huffman & Chapman

2009). The effect of parasitic infections on primates at the population, group, or individual level may vary dramatically on numerous factors related to the parasite species present, including its virulence, and the degree of infection (May & Anderson 1978), or because of variations in the susceptibility of the host due to their natural immunological responses to the infections, their demographics, habitat utilization, and diet (Stuart et al.

1998; Hahn et al. 2003; Chapman et al. 2005; Nunn & Dokey 2006; Weyher et al. 2006;

MacIntosh et al. 2010).

Parasite infections pose a serious risk to the fitness of an individual, and as a result, primates may respond to potential parasite infections through various behavioral mechanisms, to either avoid or control infections. Proposed adaptations to decrease the likelihood of future infections in primates are numerous, and include modification of ranging behaviors, and reductions in group size or subgroup formation (Freeland 1976,

1979; Chapman et al. 2009b). Several non-nutritional foraging behaviors have also been best explained as behavioral responces to parasite infections. For instance, to control existing intestinal parasites, Chimpanzees have been reported to consume whole plant leaves with rough or bristly surfaces without chewing, a behavior which has been correlated to an increased expulsion of several nematode parasites (Wrangham 1995;

Huffman et al. 1996; Huffman & Caton 2001; Huffman & Chapman 2009; McLennan &

Huffman 2012). In addition, the consumption of certain bitter plant foods, likely selected for their anti-parasitic chemical compounds, has also been attributed to parasite control in

primates (Ohigashi et al. 1994; Huffman 1997) and a variety of other, non-primate,

organisms (Poulin 1995; Lozano 1998).

Environmental factors have been identified as one of the most important causes of variation in parasitic infections. Primates living in environments with higher moisture or

humidity, such as lowland rainforests or riverine habitats, are typically found to have

higher parasite prevalence and species richness (Freeland 1976; Stuart & Strier 1995;

Stuart et al. 1998). Because moisture declines along an altitudinal gradient, primates living within areas of steep topography provide excellent natural systems to investigate the dynamics of host-parasite interactions. A study of baboons (Papio cynocephalus) in montane (>1800m asl.) and coastal lowland (<200m asl.) habitats found declines in the diversity of gastrointestinal parasites to negatively correlate to elevation (Appleton &

Henzi 1993). However, contrary to predictions based solely on environmental variations, the intensity of the infections (in terms of egg output) in this population of baboons was significantly higher within the montane habitats. The authors attributed this unexpected result to either an immunosuppression response caused by nutritional stress, as has been found in other species (Chapman et al. 2006), or to variations in foraging strategies, especially an increased consumption of insects in the montane habitat, which might promote the consumption of intermediate parasite hosts (Appleton & Henzi 1993).

As indicated by the differences found in montane and lowland baboons, diet and

foraging strategies also have a strong influence on parasitic infections. Frugivores

typically have higher dietary diversity than folivores, and thus are likely to ingest a

greater number of intermediary parasite hosts, and thus host a greater number of parasite

taxa (Dunn 1968; Nunn & Altizer 2006). Conversely, folivores consume a higher volume

of food, and are predicted to ingest an increased number of parasites from fewer species,

leading to lower species richness, but increased infection loads (Nunn et al. 2003).

However, considerable variation exists between primates at the individual, population,

and species levels, that have yet to be explained, and more work is needed to identify

broader trends in the role of diet in parasite infections (Chapman et al. 2009a).

We examined the gastrointestinal parasite diversity and prevalence in

unhabituated, free ranging Bioko Island drills (Mandrillus leucophaeus poensis) within

the Gran Caldera de Luba and Southern Highlands Reserve. The drill is considered to be

one of the most threatened primates in Africa (Grubb et al. 2003), and despite the

potential importance of parasitic infections on species conservation, information on the

gastrointestinal parasites of drills is extremely limited. An early, but extensive checklist

of the “baboon” parasites by Myers and Kuntz (1965) included both drills and their

congener, the mandrill (Mandrillus sphinx), however this was a preliminary compilation

of qualitative information from a variety of sources, much of which was from captive

individuals and experimental data, limiting its comparability to natural populations.

Considerable data are available concerning the gastrointestinal parasites of wild

baboons (Papio sp.) (e.g.: Appleton, Henzi, Whiten, & Byrne, 1986; Bezjian, Gillespie,

Chapman, & Greiner, 2008; Hahn, Proulx, Muruthi, Alberts, & Altmann, 2003; R. E.

Kuntz & Myers, 1967; R. Kuntz & Myers, 1966; Ryan & Brashares, 2012), which

indicate that drills have the potential to host a rather large number of parasite species,

including numerous zoonotic and anthropozoonotic taxa. Quantitative surveys of the gastrointestinal parasite communities of Mandrillus populations, which we will compare to in this study, have been performed on semi-captive mainland drills (M. l. leucophaeus)

of the Afi Mountain Drill Rehabilitation and Breeding Center, in Cross River State,

Nigeria (Mbaya & Udendeye 2011), and mandrills of the Centre International de

Recherches Médicales, Franceville (CIRMF), Gabon (Setchell et al. 2007). No such information is available on wild Mandrillus spp.

Bioko Island drills are limited to a 510 km2 protected area, the Gran Caldera and

Southern Highlands Scientific Reserve (GCSH), which makes up the southern third of the

island the southern third of the island. Within this area, drills are encountered from the

coastal lowland forests (0-900 m asl) up to the montane forests, at least 1,200 m asl.

Because of low fruit availability in montane forests, drills in this habitat were found to

have a primarily folivorous diet, compared to the primarily frugivorous diet found in the

lowlands (see Chapter 2: Feeding Ecology). As a consequence of the nutritional

differences between these diets, food intake of drills in the Caldera was significantly

higher than those in lowland forests. The average total dry volume of fecal samples

within the montane forest site, Caldera (mean: 262.3 cm2; range: 27-663; SD + 21.0) was

significantly greater (by 3.7 times), than samples collected at the lowland forest site,

Moaba Playa samples (mean: 71.8 cm2; range: 8-145; SD + 9.2). A significant positive

correlation was found between the volume of fiber and the total sample volume,

indicating that the total amount of food consumed was a factor of the level of folivory.

The objectives of this study were to: 1) determine if variations in either the environmental conditions or drill dietary strategies that exist between montane and lowland forests, impact the species richness, prevalence, or diversity of the infections, 2) provide baseline parasitological data on wild members of the Mandrillus genus, 3)

compare these results with the better studied Papionins. Finally, we discuss the implications of our findings to drill conservation and human health on Bioko Island.

Methods

Study Site

Bioko Island is a steeply volcanic, continental shelf island, separated from

mainland Cameroon by a shallow, 37 km wide channel created by rising sea levels

approximately 12,000 years ago. The island consists of three peaks: Pico Basile 3008 m

asl in the north and two smaller peaks, Gran Caldera de Luba (2260 m asl) and Pico Biao

(2009 m asl) in the south. The primate species inhabiting the island today are assumed to

be a subset of those found in the coastal rainforests at the time of separation. Each of

Bioko’s seven anthropoid species are represented on the immediately adjacent mainland,

but other primate species (including chimpanzees, lowland gorillas, mangabeys, mona

monkeys, etc.) have not been recorded on the island.

Drill fecal samples for parasite analysis were collected from two locations within

the GCSH. The higher elevation site “Caldera” was located within the crater of the

second largest peak on the island, the Gran Caldera de Luba. The low elevation site,

“Moaba Playa” encompassed the coastal lowland forests, and included a roughly five km

length of coastal lowland forest from the Moaba Playa camp, east to the southeastern

point of the island, Punta Dolores (Figure 1.4: Chapter 1). Compared to the lowland

forests on Bioko Island, the montane forests are characterized by lower average annual

rainfall (3.5-7.0 m vs >10m) and plant diversity, as well as an increased density of

understory vegetation (Teran 1962; Juste & Perez Del Val 1995; Lenton et al. 2000).

In addition to these ecological differences between the two forest types,

differences also exist in the use of these areas by humans. While there are no farms or

villages in the immediate vicinity of either collection site, the village of Ureca is roughly

8 km from where samples were collected at Moaba Playa, and residents sporadically

travel through trails in the area. Several clandestine bushmeat hunting camps are located

in the forest and coastline near Moaba Playa, and for five months each year, a team of

approximately 10 sea turtle researchers continuously reside at the field research camp

adjacent to Rio Moaba. In addition, a series of watersheds that feed into several rivers in

the southeastern corner of Bioko Island, including Rio Moaba, Rio Biadji, begin in the

land near the village of Moka, which are used for subsistence and commercial agriculture and cattle pastures. Although no analyses of the water downstream of these tributaries

have been performed, it is likely that the runoff from the residents of Moka and the local

farming activities travel through to the lowland rivers at Moaba Playa. In contrast, the

Caldera is at a higher elevation than any village on the island, and is accessible by a single, relatively difficult route, limiting its use for activities of both local residents and research.

Subjects

Samples were collected from wild, unhabituated groups of Bioko Island drills living in the Caldera and Moaba Playa. The fecal samples utilized in this study were collected as part of an ongoing analysis of the dietary strategies of the Bioko Island drill in relation to altitudinal variations in resource availability (see Chapter 2: Feeding

Ecology). No information is available on the home range size or daily travel distance of

drills on Bioko Island. However, it is highly unlikely that individuals from either site

would move to the other within the study periods. This is supported by daily observations

of the groups in Moaba Playa, which indicate that the same individuals utilized this

location from January to March (Justin Jay, personal communication, September 20,

2013). Estimates of the home ranges of mainland drills (10-20 km2; Astaras, 2009) and

mandrills (11-29 km2; J Hoshino, 1985; Jiro Hoshino, Mori, Kudo, & Kawai, 1984) also suggest that the 20 km separation between the Caldera and Moaba Playa is well outside

of the maximum home range of individuals at either site.

Sample Collection

We collected a total of 29 fecal samples from the Gran Caldera and 22 samples

from Moaba Playa sites samples during the first two months of 2011 and 2012,

respectively. Samples were collected opportunistically throughout the day, from

unknown individuals while surveying pre-existing census trails and rivers and subsequent

to individual or group encounters. Fecal samples were only collected if they were less

than one day old, and were not significantly damaged by insects. Samples collected in

2011 were stored in 10% formalin solution. To allow for further molecular

characterization of the present parasites, the samples from 2012 were stored in a 2.5%

aqueous (w/v) potassium dichromate (K2Cr2O7) solution. Samples of 2 g were placed in

a 15 ml cylindrical tube, and fixative was added until the total volume reached

approximately 14 ml. The sample was shaken vigorously until the bolus was broken and an equal consistency was achieved. Once back in the United States, the fecal samples were shipped to the Division of Parasitic Diseases & Malaria, Center for Disease Control,

where formal ethyl-acetate (FE) concentration of the stool samples was performed and

the sediment was examined by fluorescent and bright field microscopy to detect and

identify intestinal parasites.

Data Analysis

The number of samples positive for each parasite species was compared between

the Caldera and Playa Moaba sites using Fishers Exact tests. Differences in the mean

number of species infecting individuals at each site were compared using Welch’s t test.

To determine if there were any relationships between the presence and absence of each

parasite species in an individual and their diet generalized linear models (GLM) with

binomial distributions (logit linked function) were used. Separate models were run for

each parasite species, and explanatory variables used in the model included the site (as a

factor), and the weight (g) and volume (cm2) values of the fruit and fiber remains within

each fecal sample (fecal analysis methods outlined in detail within Chapter 2: Feeding

Ecology). These explanatory variables were also used to test if the parasite species richness of each individual was related to diet, using a GLM with poisson distribution

(log linked function). Correlations between the dietary variables in each model were assessed using the variance inflation factor (vif). None of the variables were found to have vif values over 6.0, therefor all dietary variables were retained in the full models.

Stepwise regressions were performed on the full models using AIC as the selection criteria, to limit the model to the most significant explanatory variables. The full and reduced models were compared using a likelihood ratio test.

All analyses were performed using R statistical software v 2.15.0 (R, 2012).

Significance tests were two-tailed with the significance level set at 0.05.

Results

Parasite Prevalences by Site

We collected 29 samples within the Caldera and 22 samples from Moaba Playa.

With the exception of a single sample collected at Moaba from viscous, loose stool, all

were from well-formed bolus. In both locations a total of six parasite morphotypes were

identified, three of which we were able to identify to the species level (Table 3.1). We

recovered two Protozoan species, Balantidium coli and Cyclospora papionis, and four

Nematode worms, including Trichuris trichiura, Subulura sp., and those from the order

Strongylida (Strongyle) and Rhabditida (lungworms), which we were unable to identify

further due to an absence of adult worms.

The mean species richness of individuals in the Caldera (1.52 + 0.8 SD, range=0-

3, median=1) was slightly higher than that at Moaba Playa (1.36 + 1.1 SD, range=0-4,

median=1), however this difference was not significant (t(36.5)=0.56, p=0.58) (Figure

3.1). Within the Caldera 93.1% of samples were found positive for infection by at least

one parasite species, and 77.3% were infected at Moaba Playa (Figure 3.2). No

significant differences between sites were found in the infection prevalences of any

species.

Parasitic Infections and Diet

No relationship was found between the parasite richness and the site or

composition of the fecal sample. Analyses of the relationships between the presence of

each parasite and the collection site and dietary composition of a fecal sample were

performed on all taxa except the lungworm and Subulura sp., as they were both found

only in the Caldera and at extremely low frequencies. Stepwise regressions of the full

models were largely unsuccessful at explaining the variance in the infection presence for all parasites except the Strongyles. The reduced model for Strongyles with the highest

AIC included both the site and fiber weight, with a marginally significant effect of site

(z=2.22, p = 0.03), and no significance in the effect of fiber. Based on these results, a

second model was fit which included an interaction between fiber and site weight, which

resulted in a higher significance value for the effect of site (z=2.6, p<0.01). However,

there was little difference between the null and residual deviance of the model, and the residuals only explained 15% of the total deviance. Subsequent comparisons of this reduced model with both the full model and the model fit only with the site variable were not significant for any of the species.

Discussion

Despite our expectations, no significant differences were found between the gastrointestinal parasite infections of Bioko Island drills living in montane and lowland forests. There was also no clear relationship between the consumption of fruit or fiber, or the total volume of the fecal sample, on the presence or richness of parasites in a fecal sample. These results are puzzling, as the richness, prevalence, and community of parasitic infections have been shown to vary considerably due to a myriad of behavioral and environmental factors (Snaith & Chapman 2008; Chapman et al. 2009a). The lowland forests in southern Bioko Island have relatively high rainfall and humidity, and a

large number of riparian systems, which are expected to increased richness and

prevalences of infections of primates in this habitat (Stuart & Strier 1995; Stuart et al.

1998; Bezjian et al. 2008). In addition, dietary differences between drills living in these

two forest types also increase the likelihood that their gastrointestinal parasite infections would differ considerably (Dunn 1968; Nunn et al. 2003; Nunn & Altizer 2006). Yet we found no evidence of variation in the species richness or prevalence, or the community of parasites in between drills in these habitats. These results are consistent with the variability in parasite infections found throughout the primate literature, and despite considerable attention by the research community, broad trends in the factors driving such variations have been difficult to identify (Chapman et al. 2009a; Benavides et al.

2012).

The simplest explanation for the lack of difference we found is that drills may move between montane and lowland forests at a rate faster than the turnover of these parasites within their systems. Astaras (2009) estimated that the home range of drills in

Korup NP, Cameroon, was between 10 to 20 km and their average travel distance was roughly 2.9 km/day. These estimates were similar to the home range size predictions (5-

28.5 km2) and daily travel estimates (2.5-4.5 km/day) of mandrills in the lowland forests of Campo Reserve, Cameroon, made by Hoshino (1984, 1985). However, reports of

mandrills in the gallery-forest mosaic landscapes of Lope, Gabon, have estimated that

they may have home ranges up to 182 km2 (White 2007), suggesting that the habitat type

occupied may significantly impact the ranging behavior of Mandrillus spp. Without

knowledge of the movement of drills on Bioko Island, there is no way to rule out the

possibility that the parasites drills acquire at low elevations are carried into the montane

forests during sporadic or regular travel events. Such movement may account for the lack of difference in the samples collected between the forest types.

An alternative explanation may be that certain human activities, specifically those related to the efforts to eliminate onchocerciasis (river-blindness) from Bioko Island, have modified the parasite community or host-parasite dynamics on the island.

Onchocerciasis is a disease caused by infections of the nematode Onchocerca volvulus

(Superfamily: Filarioidea), which can cause blindness and severe skin disease, including

lymphadenitis and elephantiasis, in humans (WHO 2013). In a concerted effort to

eliminate onchocerciasis from Bioko Island, the World Health Organization (WHO) carried out a program between 1995 and 2005, aimed at eradicating the vector for O. volvulus, an endemic form of blackfly (Simulium yahense), from the island. Over three application periods (2001, 2003, and 2005), 87 rivers across the island, including all those in the vicinity of Moaba Playa, were treated with the larvicide temephos

(phosphorothioate). Rivers were treated at 850 dosing points at elevations less than 500 m asl., leaving the Gran Caldera untreated (Traoré et al. 2009).

Temephos is highly effective in eliminating blackflies and mosquitoes, and is considered the least lethal larvicide to non-target organisms (Amakye 2005). Studies on the toxicity of temephos, which have primarily been performed on vertebrates, have found it to have adverse effects in relatively few species (Brown 2003). However, it has been indicated as acutely toxic at relatively low doses in numerous invertebrates, including insects (e.g. Diptera, Homoptera, and Hymenoptera), snails (Basommatophora and Gastropoda), and arthropods (e.g. Daphnia spp., Decapods, ) (Brown 2003; WHO

2011). On Bioko Island, treatment of temephos in Rio Musola was found to decrease the

population density of aquatic macroinvertebrates by 31.7% within 24 hours (Amakye

2005). Certain species, such as larvae of the caddisfly, Cheumatopsyche falcifera, were

completely lost from the river following treatment. Very little is known concerning the

potential impact this larvicide may have had directly on protozoan or nematode parasites,

or indirectly on other host vectors. It is possible that the use of temephos on Bioko Island may have modified the structure or dynamics of the island’s parasite community, resulting in similar infections of drills across habitats and elevation ranges.

Another factor that may impact the parasite community and infections in Bioko

Island drills is its unique climate. The southern lowland forest of Bioko Island is one of the wettest places in the world, and at over 10 m annually, it has the highest average rainfall recorded in Africa (Nosti 1942; Leroux 2001). Although moisture is often thought to support parasite infections, and certain species may only be found during the wet species, the prevalence of others have been shown to increase during the dry season, or remain stable throughout the year (Pettifer 1984; Bezjian et al. 2008; Trejo-Macias &

Estrada 2012; Benavides et al. 2012). Species may have unique lifecycles or morphological characteristics which vary their success in certain conditions, such as,

Oesophagostomum sp., which can arrest their development as larvae in response to the

threat of desiccation during the dry season (Pettifer 1984).

The high rainfall on Bioko Island may exert a strong selective pressure on the

endemic parasite community. For example, in a test of the effect of rainfall on

contaminated feces defecated by white-eyed mangabeys (Cercocebus albigena) onto

leaves and branches, rains were found to wash parasitic protozoans from all exposed

surfaces within 24 hours (Freeland 1980). Parasites in the feces that had been covered,

and not washed away, were killed by fungus within this time period. Under dry

conditions, however, parasites were present and alive at the 24 hour mark. It is reasonable

to suggest that, in this case, species with traits which increase the likelihood of being

washed away or killed by fungus before transmission to a host may be selected against on

Bioko Island.

Parasites of the Papionin

To our knowledge, the work presented here represents the first study of the gastrointestinal parasites of any Mandrillus spp. in the wild. Parasitological data have been reported on both drills and mandrills at the Afi Mountain Primate Conservation

Center, in Calabar, Nigeria, and mandrills the Centre International de Recherches

Médicales, in Franceville, (CIRMF) Gabon. However, as these studies were performed on individuals in captivity, they may not accurately represent the infections of drills in their natural habitats. The reported infections of free-ranging drills at Afi Mountain are considerably different than those of Bioko drills. Mbaya and Udendeye (2011) found gastrointestinal parasites to be present in only 35.3% of individuals, considerably lower than the average of either Moaba Playa (77%) or the Caldera (93%). Bioko drills were also infected by a completely different parasite community, limited to three nematodes,

Ascaris lumbricoides, Ancylostoma duodenale, and Strongyleoides stercoralis. Parasite infections of mandrills at this location were similar to those of the drill. Only 50% of individuals were infected with intestinal parasites, and by only two species, both found in the drills, A. lumbricoides, A. duodenale (Mbaya & Udendeye 2011).

Mandrills at CIRMF were found to host at least seven different gastrointestinal

parasite taxa, including three amebic protozoans, Entamoeba coli, E. histolytica/dispar

complex, and Endolimax nana, and four nematodes, including Mammomonogamus sp.,

the eggs of several unidentified taxa, and two found in Bioko drills, Balantidium coli and

Trichuris sp. The average prevalence of the taxa found in these mandrills (58%) was nearly double that of those collected on Bioko (31.3%). With respect to the shared parasite taxa, Balantidium coli prevalence was higher in mandrills (80% vs Caldera:

62%; Moaba Playa: 50%), however Trichuris sp., which was found in 38% the feces on

Bioko, was only in 1% of samples in these semi-captive mandrills. Without quantitative information on wild mandrill parasite infections, there is no way to determine if the relatively high parasite richness and prevalences found in CIRMF mandrills are a result of factors related to captivity, or natural behavioral or ecological differences between the species.

Compared to reports of other Papionins in completely wild conditions, we found

Bioko Island drills to host a relatively low richness and prevalence of parasite species, falling within the values of other species. For instance, Chacma baboons (Papio ursinus)

living on the edge of the Namib Desert in Tsaobis Leopard Park, Namibia, were infected

by a high number of intestinal parasite taxa (11), including three species found in over

two thirds of the population. However the average infection rate of all the parasites was

low (28.8%) (Benavides et al. 2012). Olive baboons (Papio anubis) in Kibale National

Park, Uganda, which might be expected to have much higher parasite infections than the

Chacma baboons living in the arid landscape bordering the Namib Desert, were infected

by fewer taxa (8), at a nearly identical average infection rate (28.5%) (Bezjian et al.

2008). Compared to either baboon species, the richness was considerably lower (6),

average prevalence among all taxa in Bioko Island drills was higher at both island

locations (Caldera = 30.3, range = 3.4 - 62.1%; Moaba Playa= 39.8%, range = 27.3 –

50.0%).

Although neither of these studies indicated the proportion of individuals infected

with at least one parasite, 85% of Olive baboons were infected by Oesophagostomum sp., and 77.5% of Chacma baboons were infected with Streptopharagus pigmentatus. This indicates that the proportion of individuals with at least one species is at least as high as that of Bioko Island drills (85%), and likely to be much higher.

Comparison of these results with wild crested mangabeys (Cercocebus galeritus), which were infected by 13 parasite species (including ten nematodes and three protozoans), again show the richness of drill parasites to be relatively low (Mbora &

Munene 2006). However, the prevalence of infections in drills was relatively high, as parasites were only detected in 57.3% of the mangabeys population (Mbora & Munene

2006). In this study, prevalences were found to be low among all taxa (11 of 13 were in

10% or less of the study population), and the average of all 13 taxa was only 5.3%. In contrast, the average infection rate of parasites in Bioko Island drills was 31.3%, and each of the taxa, except Subulura sp., were present in over 23% of the fecal samples analyzed. Infections of Trichuris trichiura was also found in crested mangabeys, however its prevalence (15.5%) was roughly half that found in this study at Moaba Playa (27.3%), and less than a third of the rate in the Caldera (48.3%).

The recovery of Cyclospora papionis was unexpected, as this is the first report of

Cyclospora in monkeys from West Africa, and extends the host range to another different

and distinct group of primates. Eberhard et al. (1999) described three new species of

Cyclospora from Ethiopian monkeys in 1999, and noted close morphologic and

molecular similarity between these three species and C. cayetanensis of humans. A

subsequent survey for cyclosporiasis in Kenyan primates confirmed the widespread

distribution of these three species, Cyclospora papionis in baboons, Cyclospora

cercopitheci in vervets, and Cyclospora colobi in black and white colobus monkeys in

East Africa, and the very marked host specificity, even in areas where the three species of

primates overlapped (Eberhard et al. 2001). Since those initial studies, there has been

one additional report of Cyclospora in vervets and baboons in Ethiopia (Legesse & Erko

2004), a reference to the presence of the parasite in hamadryas baboons in a zoological

garden in Spain (Cordón & Prados 2008) , and the description of C. colobi-like

organisms in Snub-nosed Golden colobus monkeys in northwestern China (Zhao et al.

2013).

It is currently unclear how C. papionis was transmitted to drills on Bioko Island.

Drills do not overlap with olive baboons in range or habitat type, indicating that some

third, unknown host may have been involved (Eberhard et al. n.d.). Red-capped

mangabeys (Cercocebus torquatus) are a probable suspect, as they are commonly found

with drills on the mainland (Astaras et al. 2011), and may have some range overlap with

olive baboons near the Nigeria-Benin boarder. The phylogenetic proximity and niche

similarity of mangabeys suggests that they would also be susceptible to C. papionis.

However additional work is needed to determine if this is true.

Implications for Drill Conservation and Human Health

Despite their distinction as an IUCN Endangered Species (IUCN 2013), drills are

regularly hunted and consumed as a bushmeat item throughout their range (Steiner et al.

2003; Albrechtsen et al. 2007; Morgan et al. 2013). These activities not only threaten

drills, but they also increase the high risk of the transmission of zoonotic parasites to the

local human population, particularly as each of the five parasites we found are potential

zoonotic agents (Noble & Noble 1971; Stoner 1996; Eberhard et al. 2001; Bezjian et al.

2008). The presence of two species infecting drills, Trichuris trichiura and Balantidium

coli, have also been confirmed in humans on Bioko Island (Roche & Benito 1999).

While B. coli was found at a low prevalence (<1%), T. trichiura was in up to 36.4% of

the study population. Although most of these infections are likely due to poor water

quality and sanitary conditions, it does suggest that zoonotic or anthropozoonotic

transmissions are possible.

The threat of zoonotic transmission from bushmeat activities on Bioko Island is

particularly concerning in light of the high rate of HIV infection, poor nutritional and

health status, and the immunodeficiency caused by these conditions (Chandra 1997;

Roche & Benito 1999; Roka et al. 2012). Parasites can both take advantage of such

situations, and exacerbate them significantly (Noble & Noble 1971). A study on HIV

positive patients on Bioko Island found that they had a 59.9% increased likelihood of

parasitism by T. trichiura, compared to HIV negative patients (Roka et al. 2012). Another study found that compared to WHO standards, 35.2% of children on Bioko Island under five years old had stunted growth, 10.6% were underweight for their age, and 2.8 were wasting, indicating acute starvation or disease (Custodio et al. 2008). These results

suggest that a high proportion of the human population on Bioko may have increased

susceptibility to parasites and other infectious zoonotic diseases carried by primates,

including hepatitis, Herpes B, tuberculosis, and interactions should be avoided.

Conclusions

Contrary to expectations, we found no evidence that the environmental or dietary

differences existing between the drills in montane and lowland forests impacted either the

richness or prevalence of infections. We provide three potential explanations for their

similarity: 1) altitudinal ranging patterns of drills may expose individuals to similar parasite species and infection prevalences throughout the Gran Caldera and Southern

Highlands Scientific Reserve, 2) treatment of the rivers below 500 m asl with larvicide

may impact the species richness, presence, or host dynamics, resulting in infections

similar to montane forests, or 3) the climate on Bioko Island may select against certain

parasites, limiting the species to those capable of withstanding high rainfall throughout

the year. There is no way to confirm or reject any of these hypotheses with the data

currently available, however, each present a plausible scenario that can readily be tested

in future studies.

Given the dissimilarity in the infections of wild Bioko Island drills and both drills

and mandrills in captive care, and the phylogenetic proximity of these primates, future

studies of the gastrointestinal parasites of these animals may be able to provide unique

insight into the effects of captivity on primate parasites. Comparisons of such work with

the baseline data on Bioko Island drills presented here may also increase our

understanding of the extent to which environment or behavioral characteristics, or island

isolation vary the parasite infections of primates. Ultimately, this study provides further

support for the complete ban of all primate hunting activities on Bioko Island, as well as

restricted access to the Gran Caldera and Southern Highlands Scientific Reserve to

benefit both human and non-human primates.

Figure 3.1: Counts of the species richness of parasites found in drill fecal samples collected at the Caldera and Moaba Playa.

Figure 3.2: Frequency of parasitic infections occurring in fecal samples collected at the Caldera and Moaba Playa.

Transmission Human Parasite Morbidity/Mortality Caldera Moaba Playa Cycle, Mode Pathogen Protozoa Direct, Dysentery, vomiting, weight loss, Balantidium coli Y 18/29 (62.1%) 11/22 (50.0%) Water potentially fatal Cyclospora papionis Direct Y Diarrhea 3/29 (10.3%) 8/22 (36.4%) Nematoda Lungworm Direct Y Respiratory Damage 1/29 (3.4%) 0/29 (0%) Strongyles Direct Y Diarrhea, anemia 8/29 (27.6%) 10/22 (45.5%) Subulura sp. Direct Y Typically asymptomatic 2/29 (6.9%) 0/22 (0%) Trichuris trichiura Direct Y Typically asymptomatic 14/29 (48.3%) 6/22 (27.3%)

CHAPTER 4: GROUP SIZE, POLYSPECIFIC ASSOCIATIONS, AND HABITAT USE

Introduction

Primates have been described as the most gregarious social mammals, primarily

because of the fact that all species live in social groups (Price & Stoinski 2007; Nystrom

& Ashmore 2008). However, as is the case with most aspects of the ecology and behavior

of primates, considerable variation exists between, and within, species and populations.

The number of individuals in foraging groups, defined by Jolly (1985) as the individuals

moving and feeding together, ranges between species, from solitary individuals (e.g.

Loris tardigradus, Perodictus potto, Microcebus murinus), to those averaging over 100

individuals (Papio papio and Theropithecus gelada) (Eisenberg et al. 1972; Jolly 1985a).

However, even species that spend most of their lives solitary, including most nocturnal

prosimians and Orangutans (Pongo spp.), have been shown to form dispersed polygynous

mating groups, and often exhibit complex social behaviors (van Schaik & Van Hooff

1983; Galdikas 1988; Nekaris 2006; Price & Stoinski 2007). This remarkable diversity in

social structure and behavior across the Order has been one of the primary interests

primate research (Snaith & Chapman 2007). Such variation provides a means in which

the ecological determinants driving social organization can be investigated, and the

evolutionary pressures involved in facilitating sociality may be better understood.

Theoretical models and empirical field studies have identified numerous factors

which may have a strong influence on the size and structure of primate groups, including

predation pressure, resource defense, mate access, and genetic constraints (Snaith &

Chapman 2007; Nystrom & Ashmore 2008). However, attempts to establish broad

generalities of the causative factors involved have been relatively unsuccessful, owing to

the complexity of the interactions between these agents. One commonality that has been

found in a wide range of taxa is that the availability and distribution of foods, and the

dietary strategy of the primate can play a particularly strong role in group dynamics

(Clutton-Brock 1974; Wrangham 1980; Byrne & Whiten 1993; Chapman et al. 1995).

Diet and foraging can place both associative and dissociative pressures on the

group size and organizations of primate groups, and the variations in grouping exhibited

between and within species are a result of these competing pressures. The benefits of

large groups to foraging success can be large, including an increased ability to discover

foods (Vickery et al. 1991), defend limited resources (Wrangham 1980), or avoid

predators (Hill & Lee 1998). However the costs may also be high. The majority of the

daily activity budget of primates is spent feeding, resting, or traveling to food sources, the

rest is spent on “non-subsistence behaviors” needed to maintain intragroup relationships

(grooming, play, vocalizations, etc.) (Bartlett 2009). The time spent on these social

activities has been shown to correlate with increasing group size (Dunbar 1991).

However, in situations where food resources are scarce, individuals may decrease time spent on non-subsistence behaviors to increase their foraging time and meet their minimum dietary requirements. For example, Defler (1995) reported a significant decline

in the proportion of the daily time budget devoted to social activities, from 9% to 1%, in

woolly monkeys (Lagothrix lagotricha) during periods of fruit scarcity. Similar declines

in social activity time have been reported in white-handed gibbons (Hylobates lar) (20%

to 3%) and wedge-capped capuchins (Cebus olivaceus) (20% to 6%), in response to

seasonal declines in fruit availability (Robinson 1986; Bartlett 2009). The time required

to maintain large groups may represent a significant cost to large primate groups, and

may promote reductions under certain food availability conditions.

Another cost of large groups is that the increase in biomass requires higher food

intake. Frugivores are especially prone to such constraints, as fruiting trees in tropical

rainforests often produce fruit in spatiotemporally clumped patterns, resulting in times

and areas with little fruit available to consumers (van Schaik et al. 1993). Such patterns in

food availability are expected to increase the intraspecific and interspecific competition

between primates sharing these resources, and place a strong selective pressure against

increased group sizes (Chapman 1990a; Isbell 1991; Steenbeek & Schaik 2001; Snaith &

Chapman 2007). However a range of behavioral and ecological mechanisms have been

proposed as mechanisms to cope with reductions in food availability, and reduce the

competitive pressure on large groups.

For instance, although true migration is not found in any species of primates

(Jolly 1985a), they will commonly shift their core range use in response to food

availability (Gautier-Hion et al. 1981; van Schaik et al. 1993; Wallace 2006; Vogel et al.

2009). Collard mangabeys (Cercocebus torquatus), which form large foraging groups

(mean = 21.1 individuals), utilize subsets of their total home range in response to

seasonal shifts in fruiting tree availability (Mitani 1989). Likewise, Ganas & Robbins

(2005) found positive correlations between fruit availability and both the daily travel

distance and home range size of mountain gorillas (Gorilla beringei beringei). Instead of

shifting or expanding their home ranges, other species may respond to local scarcity of

foods by concentrating their ranging to specific habitats within their home range, such as

riparian or streamside vegetation as with collared titi monkeys (Callecebus torquatus;

Kinzey, 1977), or swamp habitats grey-cheeked mangabey (Lophocebus albigen;

Poulsen, Clark, & Smith, 2001).

Because of the considerable variation in the group size and behaviors exhibited by

primates, species and population specific data on the ecology and habitat use of primates

is particularly important to conservation in tropical forests. Primates have been found to

make up a large portion of the mammalian biomass in numerous tropical forest sites

(Eisenberg & Jr 1973; Prins & Reitsma 1989; Oates et al. 1990; White 1994; Chapman &

Onderdonk 1998), and can play a vital role in the maintenance of their forest ecosystems

through numerous mechanisms. This is particularly true of role large frugivorous

primates play in seed distribution (Chapman et al. 1995; Chapman & Onderdonk 1998;

Clark et al. 2001). The loss of large seed distributors in tropical forests can have long-

term effects on the community composition and dynamics, and ecosystem functions of a

tropical forest (Chapman & Onderdonk 1998; Fa et al. 2002; Effiom et al. 2013).

Therefore information on the ecology, habitat use, and ecological drivers of variations in

the population dynamics of primates is critical to understanding, and potentially

mitigating, the impacts that local extirpations may have on the remaining sympatric

community members.

In this study we investigated how ecological variations in the structure and

composition of montane and lowland forests impact the habitat use and group size of

Bioko Island drills (Mandrillus leucophaeus poensis). Like much of their ecology and

behavior, the group size and structure of wild drills is poorly known throughout much of

its range. Reports of group sizes reported on the mainland subspecies (M. l. leucophaeus)

range from numerous encounters of solitary males, to groups with over 400 individuals

(Gartlan 1970; Wild et al. 2005; Astaras 2009). Average group size estimates, from the

encounters deemed reliable by the authors, range from 52.3 to 93.1 individuals

(Appendix 4.1). Reports of the reliable group size estimates of mandrills (Mandrillus

sphinx) are comparable to those of drills, except in the stable aggregations of up to 845

individuals (mean=620) reported in Lopé Reserve, Gabon (Rogers et al. 1996; Abernethy

et al. 2002). On Bioko Island, Schaaf, Butynski, & Hearn (1990) reported drill groups to

range from 1 to 7 individuals, but note that in several occasions additional group

members were clearly in the vicinity, but not seen. Gonzalez-kirchner & de la Maza

(1996) commented that drill groups on the island ranged from 2 to 20 individuals;

however they give no indication of the total number of encounters, or any ancillary

information to indicate the reliability of their encounters.

Habitat use information for both drills and mandrills is primarily limited to

information on their occurrence in broad habitat types and elevation ranges. On Bioko,

drills are encountered from the coastal beaches on Bioko Island, through the lowland

forests (0-900m asl.) and into the island’s montane forests (900-1,200m asl.) (Schaaf et

al. 1990; Gonzalez-Kirchner & de la Maza 1996). On the mainland, drills can be

encountered at a much higher range, up to the montane grasslands of Mount Kupe,

Cameroon at 2000m asl (Wild et al. 2005). In both the island and mainland, drills are

regularly encountered in areas with dense understories of herbaceous vegetation,

including stands of bracken fern (Pteridium aquilinum), Aframomum spp., and the grassy

slopes of montane and alpine habitats (Sanderson 1940; Schaaf et al. 1990; Wild et al.

2005). On Bioko Island, drill are regularly encountered foraging in and along rivers, and

crustaceans have been indicated as an regular food source (Schaaf et al. 1990). Finer

scale habitat (i.e. microhabitat) use information is not available on either the drill or

mandrill.

Group sizes of wild, unhabituated primate troops have been described as often

highly inaccurate and significantly underestimated by observers (Thomas 1991). In a

study in Lopé Reservé, Gabon, Abernethy, White, & Wickings (2002), compared precise

group size counts of mandrills from video recordings to observer estimates, concluding

that observers underestimated group sizes by 40%. As these estimates were derived from

counts of aggregations of up to 845 individuals, the high margin of error reported there is

likely to be correlated to the difficulty of quickly and accurately counting such large

numbers of moving primates. However, based on these findings, Wild, Morgan, &

Dixson (2005) suggest the group size estimates they reported of drills in Bakossiland,

Cameroon, be regarded in light of an equal margin of error. A common method to decrease the potential error in group counts collected, which we employed in this study,

is to assign a reliability designation to all drill groups encountered visually during

surveys (Gartlan 1970). An example of this would be if the individual were counted

while in a sleeping tree at dusk or dawn, or while crossing a river without being aware of

the observer and no other individuals were heard calling in the area.

The objectives of this study were to determine if altitudinal variations in habitat

structure, food availability, or the diet of drills impact the group size, microhabitat use, or polyspecific associations of Bioko drills. Specifically we address four questions related to their social characteristics: (1) what is the average size and composition of drills on

Bioko Island, (2) does this differ significantly between montane and lowland forest sites,

(3) what impact does the survey method have on these estimates, (4) what is the average

rate of polyspecific association with other primate species and duikers. We also investigated the distribution of drill group encounters along survey transects to determine the microhabitat associations related to drill presence at a given location. We will put this in a broader context by comparing our results to those of better studied Papionin monkeys, and discussing the conservation implications of our findings.

Methods

Study Design and Caveats

Three different sites within the Gran Caldera and Southern Highlands Scientific

Reserve were used to address different aspects of this study (Chapter 1: Figure 1.4). Data for the group size estimates and polyspecific associations were collected at all three localities; the Caldera, Moraka Playa, and Moaba Playa. Investigation of the habitat

associations required frequently monitored survey transects, which Moaba Playa lacks,

thus habitat assessments were only performed in the Caldera and at Moraka Playa.

The tropical forests habitats of Bioko Island drills are characterized by low

visibility and dramatic altitudinal relief (Zafra-Calvo et al. 2010). Given that drills are

highly elusive, terrestrial primates, and significant hunting pressure exists throughout

their range (Astaras 2009; Morgan et al. 2013), group habituation was neither ethical nor

feasible within this study period, and much of the data on group size and structure

presented herein were collected opportunistically, on and off established BBPP survey

transects, rivers, beaches, and footpaths. Field work was restricted to the dry season

(January to March) each year from 2009 to 2012. Even then, torrential downpours that

lasted several days without substantial breaks were commonly experienced during this

study, during which surveys were not performed. Attempts at field surveys in the wet

season during the exploratory work for this study in 2008 confirmed the extremely low

visibility during the wet season (often less than 5m in completely open areas) would

make observational fieldwork infeasible during this period; thus our study was limited to

the dry season.

In addition, this study was performed as part of a broader assessment of the

ecology and behavior of the Bioko Island drill, and required significant time spent during

surveys searching for fecal samples, foraging remains, and observing drills. Transects

were rarely surveyed in adherence to the methods used to estimate group abundances or

densities (Butynski 1984; Butynski & Koster 1994; Peres 1999), thus we refrained from

making such estimates in this study. Information on the relative abundance of the primate

community members in the GCSHSR have been recently estimated by others (Cronin et

al. 2010; Cronin 2013).

Group Size, Composition, and Polyspecific Associations

For each drill group visually encountered, the number of individuals and location

were recorded, and the minimum group visual counts were marked as reliable if the

observer estimated that >80% of the group was seen, based on noise (e.g., vocalizations

and movement), and circumstances of the encounter. The number of adult males and

females, sub adult males, juvenile and infants were recorded to determine the average

group composition. The number of 2 phase grunts (2PG) from distinctly different locations during visual encounters were be included, as only adult and sub-adult males make this call, and incorporation of these data can increase the accuracy of the male

count (Gartlan 1970; Astaras 2009). No other vocalizations can be reliably used to

determine sex or age class composition.

Group size data were also collected in 2012 from observations made from within

blinds set along a river in the Moaba Playa area (exact location is withheld because of the

high threat of hunting). Blinds made of natural materials were constructed on river banks

opposite to several fruiting Ficus vogeliana trees that were known to be frequented by

drills, to collect high resolution video footage of drills for documentary and behavioral

research projects. Because group size counts were often collected while whole groups

were walking along or across the river in close proximity of the observer, and the drills

were unaware of their presence, these counts were all deemed reliable. Reliable group

size estimates from transect surveys (those performed on foot) were compared between

locations to determine if size varied as a factor of the forest type occupied. To determine

the impact of survey methodology, reliable survey estimates were compared with those

derived from blinds. Finally, reliable counts from transect surveys were compared with

the estimates from all encounters on transects (reliable + unreliable) to determine if

significant differences existed between the datasets with and without reliability

designations.

Habitat Profile

Data for the habitat assessment were collected using Trimble Nomad (900 series)

handheld computer units with built-in GPS devices, running a data collection program

developed in Cybertracker v3 software (http://www.cybertracker.co.za/). To test the

relationships between drill presence at a given location and the local conditions of the

habitat, the point quarter quadrant (aka. plotless quadrant) method was used to develop basic habitat profile along the Santo Antonio trail (5.7 km) in the Caldera, and Bajda trail

(6.0 km) at Moraka Playa (Eisenberg 1981). Relative to quadrat methods, plotless sampling techniques enable rapid assessment of ecological variables across a large area, however at the expense of fine resolution.

At points in 100 m intervals along the length of each transect, four quadrants were established using the N-S and W-E lines of a compass. The forest parameters assessed at each point included canopy closure (1-4 class scale: 1 = 0-24%, 2 = 25-49%, 3 = 50-74%, and 4 = 75-100%), visibility (1-4 class scale), and the level of human disturbance (1-4 scale). As certain species of terrestrial herbaceous vegetation (e.g., Aframomum spp.,

Costus spp.) have been identified as important food items for Bioko drills (Chapter 2), and previous reports of mandrill indicate they avoid areas with densely vegetated forest floors (Lahm 1986; Wood 2007; Kingdon et al. 2013), the dominant and secondary ground cover was estimated (1-4 class scale) in a 1 x 1 m quadrat, 2–5 m off the trail in a randomly chosen direction perpendicular to the trail (Beymer & Klopatek 1992).

Groundcover type was recorded as dirt, rock, leaf litter, fern, Aframomum sp., grass, saplings, shrub, or other terrestrial herbaceous vegetation. Within each quadrant, the tree closest to the assessment point with a diameter at breast height (DBH) greater than 10 cm was located, and the DBH (cm), estimated tree height (+ 5 m), and point-to-tree distance

(m) were recorded. The mean value for each of these measurements was calculated for

each point and used in comparisons. Finally the fruit and liana presence on each of the

four trees was recorded and the sum value of each point in the statistical analyses. The

mean point-to-tree distance for all species (di) was used to determine the tree density

(trees/hectare) at a location, using the formula: d = 1/di2 · 100.

Finally, because rivers have been identified as a potentially important foraging

habitat for drills and mandrills (Lahm 1986; Schaaf et al. 1990), the distance from each

point of the habitat assessment to the closest river was calculated. Using global

information system software ArcMAP v10 (ESRI 2013), rivers were first digitized using

a digitized basemap of Bioko Island (MAP REERENCE), the 100 m habitat assessment

points along each transect were imported, and both features were converted to shapefiles.

The Near tool within the Proximity toolset was used to calculate the distance (m) from

the closest point of any river to each 100 meter point, and analyses were performed on

the Log10 transformed values.

Data Analysis

Comparisons of the group size estimates were made through individual Kruskal-

Wallis tests, as Shapiro-Wilk normality tests indicated they did not meet the normality

assumption required of analyses of variance (ANOVA). Subsequent multiple

comparisons between the three sites were made using kruskalmc from R package

pgirmess (Giraudoux, 2012). Differences in the habitat variables between the Caldera and

Moraka Playa were made using Kruscal-Wallis tests for the continuous variables (DBH,

tree density, tree height) and individual Fishers Exact Tests for the ordinal (canopy

closure, visibility, human disturbance), discrete (mean tree fruit presence, liana presence,

and liana fruit presence), and nominal (dominant and secondary groundcover types) data.

The frequency of drill presence among all habitat assessment points was compared

between trails through a Chi-Squared test. The frequency of encounters at points where

presence was recorded at least once was calculated by dividing the number of encounters

by the total number of times the point was passed during surveys or other patrols. These frequencies were compared between transects using a Welch T-Test.

The relationships between each habitat variables and the recorded presence of drills at a given habitat assessment point were determined using a binomially distributed generalized linear model (GLM), with a logit linked function. First, a full model with each of the habitat variables (including forest type) was fitted. The influence of collinearity between the predictor variables on the model variance was assessed by calculating the variance inflation factors of the model using the VIF function from R package car (Fox & Weisberg 2011). DHB and tree height were found to be highly correlated (VIF>10), therefore independent full models were fit with these two variables.

Next, a stepwise selection procedure was performed using Akaike’s information criterion

(AIC), to limit the final model to the most significant variables (Ruiz-Labourdette et al.

2012). All analyses were performed using R statistical software v 2.15.0 (R Core

Development Team 2012). Significance tests were two-tailed with the  level set at 0.05.

Results

Group Size, Composition, and Associations

Transect surveys were performed 70 times on the Santo Antonio trail in the

Caldera, and 42 times on Badja trail at Moraka Playa. Drills were encountered 128 times on Santo Antonio Trail (1.83 groups/survey or 0.32 groups/km) and 17 times on Badja

Trail (0.41 groups/survey or 0.07 groups/km). During the transect encounters, 34 (23.8%)

were deemed reliable, the majority coming from the Caldera (25); only six were recorded

at Moraka Playa and three at Moaba Playa. A total of 38 surveys were performed in

blinds, during which drills were encountered 102 times (2.68 groups/survey). Group sizes

ranged between 1 (solitary males) and a single troop of 20 individuals, which was

repeatedly observed at Moaba Playa (Table 4.1). A group of 18 individuals was observed

one time in the Caldera in 2011, all other encounters at this montane forest site were less

than ten individuals. Solitary adult males were encountered 18 times, and subadult males

25 times, making up 13.2% and 18.4% of the reliable encounters from both survey

methods, respectively.

On average, groups were composed of 23.4% males (N = 0.88), 22.4% females (N

= 0.85), 29.3% subadult males (N = 1.1), 11.8% juveniles (N = 0.5), 6.7% infants (N =

0.3), and 6.6% unknown (N = 0.24) individuals. No significant differences existed

between the reliable encounters at the three sites in either their average group sizes or the

proportions of group member classes, thus the counts from all sites were combined to

compare with those from blinds. The mean group size from blinds (3.4, +0.3 SEM) was

significantly lower than that of the transect surveys (4.2, +0.15 SEM) (Kruskal-Wallis,

H=5.9; p = 0. 015) (Table 4.1).The proportion of males and females estimated by

transects were significantly higher than those from blinds (Male: H=13.6, p < 0. 001;

Female: H=12.8, p < 0. 001) (Figure 4.1). No other differences in the proportion of sex/age classes were significant. The mean group size estimate of the reliable transect surveys (4.2) was significantly higher than the combined dataset containing both reliable and unreliable data (2.0) (Kruskal-Wallis, H=31.5, p < 0. 001), as were the increased

proportions of subadult males, juveniles, and infants (Table 4.1).

Drills were found in polyspecific associations with all other primate species

except Stampfli’s putty-nosed guenon (Cercopithecus nictitans) (Table 4.2). Associations

were recorded during 14.0% of encounters, and at all three sites. Drills were associated

most with black colobus (Colobus satanas), which were in 4.9% of encounters, followed by red colobus (Procolobus pennantii), red-eared guenon (Cercopithecus erythrotis), and

crowned guenon (C. pogonias), which were all recorded during 3.5% of drill encounters,

and Preuss’s guenon (Allochrocebus preussi) were only in 1.4% of encounters.

Habitat Assessments

Habitat assessments were performed at 46 points on the Santo Antonio trail, and

61 points on the Badja Trail. This discrepancy in surveys is due to the data from 15 points on Santo Antonio being lost due to an error with one Trimble Nomad unit. Visibility was significantly higher at Moraka Playa (Fisher’s Exact Test, p < 0.001), with 95.1% of

points having a visibility greater than 10m. The Caldera had a relatively dense

understory, with 63% of points having a visibility of less than 10m. Human disturbance

significantly lower within the Caldera (Fisher’s Exact Test, p < 0.001), however Moraka

Playa was still found to have low human disturbance levels. This difference between the

two sites is likely due to the high use of Moraka Playa for other research activities throughout the year, relative to the Caldera. Moraka Playa is the main camp for all research activities of the BBPP in the southwestern corner of Bioko Island. As a result, the trails are used with higher frequency than those in the Caldera. Mean tree fruit presence was significantly higher at Moraka Playa (Fisher’s Exact Test, p < 0.001), where 44.3% of points had at least one tree with fruit present. Only a single fruiting tree

was encountered within the Caldera (0.54% of all the trees assessed in the Montane

Forest).

Both the dominant and secondary groundcover types were found to differ

significantly between sites (Fisher’s Exact Test, each at p < 0.001). While leaf litter made

up the majority of the dominant groundcover at both locations, the points at Moraka

Playa had higher rock and “other herbaceous plants”, such as Paracostus englerianus and

Dracaena phrynioides, and the Caldera had more Aframomum sp. and Grasses. The most frequent secondary groundcover type in the Caldera was Aframomum sp., (41%) and Fern

(28.3%), whereas the Lowland Forest was Leaf Litter (30%), Rock (28%), and Saplings

(28%). Tree heights in the Caldera (mean=12.3m, +10.2 SD) were significantly shorter

than at Moraka Playa (mean=17.6m, +4.5 SD) (Kruskal-Wallis, H=102.8; p < 0.001). No

significant differences were found between the sites in their mean tree density, DBH,

canopy closure, liana presence, liana fruit presence, or river proximity.

The frequency of drill presence at each of the 100m habitat assessment points was

significantly higher along the Santo Antonio trail (0.81) within the Caldera, compared to

Badja trail (0.23) at Moraka Playa (Pearson’s Chi-squared, X2=37.04, df=1, p>0.001).

The frequency of the use of individual points where drill presence was recorded on Badja

trail (mean=7.14, SEM=+0.97, range= 5.9-17.7) was significantly higher than those used

on the Santo Antonio trail (mean=2.13, SEM=+0.19, range= 0.8-5.5) (t14.164=5.39,

p<0.001). We found the presence of drills at a given location was best explained by a

2 reduced model including the forest type (estimate=2.7489, Wald X = 5.415, p>0.001)

and river proximity (estimate=1.2139, Wald X2= 2.180, p=0.029), however this model

only explained 28.3% of the variation in drill presence.

Discussion

Group Size and Associations

Average group size estimates of Bioko Island drills are dramatically less than

those previously reported for drills or mandrills. Reliable estimates derived from both

survey methods and all sites combined result in an average group size of 3.8 individuals

per group. Even if we were to include a 40% error estimate, the maximum average would

still only be 5.3. Compared to the average group size estimate of all previous reliable group counts (except hoards) reported of drills and mandrills combined (89.7), Bioko

Island drill groups are almost 17 times smaller (Gartlan 1970; Tutin et al. 1997; Wild et al. 2005; Astaras et al. 2008). The largest group encountered during our study was only

20 individuals, and only 4% of groups had 10 individuals or more. The relatively low average and range of group sizes of Bioko Island drills remains true when compared to other genera in the Papionin tribe (Appendix 1), leading us to suggest that drills on Bioko

Island may have the smallest average foraging groups of all its members.

The high proportion of solitary males to the total number of reliable encounters

(31.6%) relative to other studies may be inflated by the fact that 88% were observed from the blinds. If the drills using the fruiting trees on the river containing the blinds were to follow similar daily paths, they would be overrepresented in the group size estimates. As no other studies of drills or mandrills have successfully used blinds, these values may not be comparable to previous reports. Solitary males were encountered in only 14.7% (N =

5) of reliable transect surveys, which is much closer to the proportion reported by previous studies of drills (Gartlan, 1970: 12.5%; Astaras et al., 2008: 6.3%; Wild et al.,

2005: 7.6%).

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Group composition was considerably different in Bioko Island drills than previous

reports of mainland Mandrillus spp. We found the ratio of adult males to the average

group size to be approximately 1:4, whereas a fairly consistent ratio of one male to 20 to

21 group members have been reported of both drills and mandrills (Gartlan 1970; Rogers

et al. 1996; Astaras 2009). At 13.9, the male ratio of mandrills visually encountered by

Hoshino (1984) was smaller than either drill study, however estimates of group

composition based on the distribution of the fresh weight of fecal samples was somewhat

closer to these values (17.8). We also found that on average, groups were composed of a

nearly equal ratio of adult males to adult females (1:0.96), and 22 groups from reliable

encounters had only one adult male and one adult female (19.0% of all non-solitary

reliable encounters). In contrast, the adult male to female ratios from reliable counts were

1:4.5 and 1:11.5 for drill groups in Korup N.P., Cameroon, and 1:10.7 for a horde of

mandrills in Lopé Reserve, Gabon (Rogers et al. 1996; Astaras 2009).

Despite the similarity between these previous studies in the ratios of males to the

total group size and to females, the social structure of Mandrillus groups has been

difficult to define. For example, mandrills in the forest-savanna mosaic habitat of Lopé,

Gabon, were found to remain in cohesive multi-male/multi-female groups of 340 to 845

individuals over a three year study period (Abernethy et al. 2002). The authors of this

study note that there was no indication that the hordes were formed by the aggregations

of smaller groups. On the other hand, mandrills in the continuous rainforest habitat of

Campo Reserve, Cameroon, were in single-male groups in 43.9% of encounters, and

multi-male groups in only 36.4% (Hoshino et al. 1984). The multi-male groups were seen

disaggregating into smaller single-male and multi-male sub-groups on five occasions, and

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the authors found a significant seasonal difference between the occurrence of single-male

and multi-male groups. These studies indicate that the group size and structure of the

Mandrillus may vary due to spatial or temporal differences. However, to our knowledge

no studies on the genus have reported single-male/single-female groups or monogamous

partnerships.

The high proportion of groups encountered in this study with only one adult male

and one female suggests that monogamy may be found in Bioko Island drills. Field

observations made during this study provide support to this conclusion, as male/female

pairs were observed on several occasions feeding on a single fruiting tree over two or

more successive days, with no indication of other individuals in the area. Monogamy is

relatively rare in primates, with less than 15% of all species thought to form such

relationships (Terborgh & Janson 1986). In Old World monkeys, the evolution of

monogamy has primarily been attributed to the combined effects of predator release and

food competition. Pressure from natural predators has frequently been associated with the

formation of large single or multispecies aggregations in primates (Gartlan & Struhsaker

1972; Peres 1993; McGraw & Bshary 2002), other mammals (Lingle 2001; Hass &

Valenzuela 2002), birds (Clark & Robertson 1979; Tellería et al. 2001), and fish

(Semeniuk & Dill 2006). Each additional individual increased the number of eyes and

ears available for surveillance of the area, thus there may be a greater chance of spotting

predators (Chapman & Chapman 1996). However, the foraging efficiency of individuals

in larger groups is expected to decrease, requiring increased time devoted to foraging

activities and larger daily travel distances or home ranges (Defler 1995; Ganas &

Robbins 2004).

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van Schaik & van Hooff (1983) note that many of the monogamous species of

Old World monkeys live in locations, including Madagascar or the Mentawai Islands off

of Sumatra, Indonesia, where the large felines that pose the greatest predatory threat are

absent. This is true of Bioko Island drills; with the exception of the African rock python

(Python sebae), none of the natural predators of drills or mandrills on the mainland

(leopards (Panthera pardus), crowned eagles (Stephanoaetus coronatus), or chimpanzees

(Pan troglodytes)) are found on Bioko Island (Harrison 2009). Therefore, we suspect that there may be little benefit to increased group sizes in Bioko Island drills, and that the

high cost of intraspecific competition has promoted smaller group, driving them towards

equal sex ratios.

The rate of polyspecific associations with drill encounters was also relatively low

compared to other estimates of drills. Astaras et al. (2011) recorded polyspecific

associations during 50% of mainland drill encounters (Table 4.2). These drills in Korup

N.P. were reported to associate with six other primate species, including three guenons found in both Korup N.P. and Bioko Island, Putty-nosed monkey (Cercopithecus nictitans), Crowned monkey (C. pogonias), and the red-eared monkey (C. erythrotis).

However, the average proportion of associations among these three species was 29% less on Bioko than Korup, and drills were never found in association with C. nictitans, which were in 39% of Korup drill encounters (Astaras et al. 2011). Mainland drills were also found with Red-capped mangabeys (Cercocebus torquatus) in 25% all encounters

(N=44). These mangabeys have similar ecological and behavioral characteristics to drills, being highly frugivorous omnivores with primarily terrestrial movement and foraging strategies, and share unique morphological adaptations for finding and consuming hard

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nuts and seeds within the leaf litter (Fleagle & McGraw 1999). Thus, they are more likely

to be food competitors with drills. Astaras et al. (2011) found that the frequency of these associations were significantly different than would be expected solely by random

chance. Based on field observations, he suggested that the associations between

mangabeys and drills may represent a predator avoidance strategy, as has been found in

other species (Hill & Lee 1998).

The rate and nature of polyspecific associations we encountered were more in line

with the observations of Gartlan & Struhsaker (1972) from Cameroon, who noted that,

”associations including drills were both infrequent and transient”, occurring only in 2%

of their encounters. Their observations indicated that most encounters of drills with other

species were a consequence of drills traveling through an area in which the other species

were occupying. This was the case with many of the encounters reported here, drills were

observed foraging within the crown of single trees with Cercopithecus erythrotis, C.

pogonias, and Procolobus pennantii for up to 15 minutes. Previous reports of

associations of both drills and mandrills with other species all indicate them to be

relatively uneventful occurrences (Gartlan & Struhsaker 1972; Sabater Pi 1972; Astaras

et al. 2011), and we found this to be true for all but one observation on Bioko Island.

During a patrol on the Santo Antonio Trail in the Caldera on February 19, 2010,

highly aggressive behavior was observed from drills towards a troop of black colobus.

The colobus group (composed of at least two males, one female with infant, and two of

unknown sex classes) had been observed foraging within two adjacent trees that were

isolated at the edge of a large patch of bracken fern and Aframomum spp., at a vertical

height of approximately 3 -5m for at least 15 minutes. At this point a small group of drills

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(only 1 male and 1 female were seen, however vocalizations indicate that additional

members were nearby) was heard approaching the area. The drills were obscured by the

high vegetation (~3m), and moved within 15m of the female and infant colobus, at which

point the large male drill charged them. The female colobus grabbed her infant and the entire group rapidly climbed up to ~15 m, where they were able to access the contiguous canopy. The male drill pursued the female and infant colobus until his large size restricted him from climbing further, at which point he returned to ground level. For the

duration of the episode (roughly one minute), both the drill and colobus groups created a

cacophony of alarm vocalizations that had never previously or subsequently been heard

by the observers. To our knowledge, this is the first published record of such high

aggression of wild drills towards other primates.

Large aggregates of conspecific and polyspecific groups require higher availability of food resources, and often necessitate a higher daily travel distance to meet their nutritional needs (Terborgh 1983; Chapman 1990b). While increased group size may benefit the individual foraging efficiency of group members, because of the group’s greater ability to find patchy resources (Gartlan & Struhsaker 1972; Gautier-Hion et al.

1983), this is not always the case. In a study of polyspecific associations of a primate

community (including mandrills and red-capped mangabeys) in Campo Reserve,

Cameroon, Mitani (1991) argued that the increased foraging efficiency resulting from

larger groups may benefit larger bodied primates less because of the dietary and spatial

requirements associated with an increased foraging biomass.

On Bioko Island they are to restricted an area of less than 500 km2 that contains some of the highest recorded primate densities in all of Africa (Butynski & Koster 1994).

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Recent surveys performed in this area have reported primate encounters at an average

rate of approximately 1.5 groups per km (Cronin et al. 2010), which is three times the

rate in Korup National Park, Cameroon (0.47) (Linder & Oates 2011). Each of the six

species that are sympatric with drills on Bioko Island consume some amount of fruit, and

therefore represent potential food competitors for the drill. Given that drills are one of the

largest monkeys in the world, with males weight over 30 kg on average (Hill 1970), the

small foraging group size and low frequency of polyspecific associations may reflect

strategies to avoid intraspecific and interspecific competition. This may also explain the

dietary differences observed in the montane forests (Chapter 2), where in response to low

fruit availability, drills switch to a primarily folivorous diet never previously reported in

drills on the mainland.

There is also strong evidence to suggest that human hunting activities may have

further promoted the reductions in group sizes and rate of polyspecific associations, and

high number of single-male/single-female pairs observed. Annie Gautier-Hion, Quris, &

Gautier (1983) noted that groups of moustached guenons (Cercopithecus cephus) that

were excessively pursued by researchers would disassociate into smaller groups, and then

find dense foliage to conceal themselves in silence. Tilson (1977) found that the

monogomous groups of Simakobu monkeys (Nasalis concolor) on Siberut Island, one of

the four Mentawai Islands off Sumatra, Indonesia, also employ such cryptic behaviors to

evade human predation, and suggested that crypsis may represent the best strategy to

avoid the unique long-range hunting ability of humans. Watanabe (1981) found that

variations in the occurrence of monogamous and polygamous groups of these Simakobu

monkeys were best explained by bow and arrow hunting by humans, supporting Tilson’s

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findings. Polygynous groups were only found in an area of the island without historic

human hunting activities, were less likely to flee from observers, and individuals often

emitted alarm chirps when approached (Watanabe 1981). In contrast, the monogamous

groups found in heavily hunted areas rarely vocalized, and either immediately dropped to

the ground to flee or concealed themselves in the canopy when approached by observers.

Although numerous measures have been taken over the past 30 years to regulate

bushmeat hunting on Bioko Island (including the formation of two large protected areas,

Pico Basilé National Park and the Gran Caldera and Southern Highlands Scientific

Reserve, and the passage of numerous legislative acts), a lack of enforcement has resulted

in intense hunting pressure throughout the island. Surveys of the primary bushmeat

market on in the capital city, Malabo, performed from 1997 to 2010 recorded the sale of

over 34,000 monkeys, and found the contemporary average rate of carcasses sold each

day (17.77 carc day-1) to be 4.2 times higher than at the start of the study (4.26). (Cronin et al. 2010). Currently, the number of drill carcasses being sold each market day is estimated to be approximately 5.6 (Cronin, pers. comm.). Drills are targeted by bushmeat hunters because of the relatively high profitability resulting from their large body, and they may be particularly susceptible to shotgun hunting, as they are easily tracked and tree forced into trees by dogs, at which point hunters can kill a large number individuals in the group (Gadsby & Jenkins 1997). During this study, drills often employed the same tactics observed of the Simakobu monkeys on Siberut Island; once human presence was detected, groups typically ran immediately, if they were in a feeding tree they often dropped to the ground (falls of ~8 m were observed on several occasions), and ceased vocalizations. It is likely that this intense hunting pressure is exerting a strong pressure

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against large group size in drills, as was found in the Simakobu monkeys (Watanabe

1981).

Habitat use

The distribution and quality of food resources in an area is expected to have a

direct impact on the size of either the foraging group size or the foraging area used by

primates (Waser 1977). Drills in the Gran Caldera have a diet primarily composed of

terrestrial herbaceous vegetation (viz., ferns, Aframomum spp., grasses, Costus spp.), which are low in quality, but abundantly distributed throughout the montane habitat (47%

of points versus 9.8% in the lowland forest). Considering that we found no difference

between the group sizes between drills in lowland and montane forests, we should expect

them to use their habitat in a considerably different manner than the primarily

frugivorous drills in lowland forests. The significant difference in the frequency of drill

presence along the Santo Antonio and Badja trails supports this hypothesis. Drill

presence was observed at 81% of the points along the Santo Antonio trail, and the largest

stretch of trail without drill presence was only 200m. In contrast, drills were only

encountered in fifth of the total points on Badja Trail, and a stretch of 2.5km (41% of the

total transect distance) was completely devoid of encounters. However, as indicated by

the significant difference in the frequency at which individual points were occupied,

drills at Moraka Playa seem to have a higher fidelity to certain areas along Bajda Trail,

whereas drills in the Caldera seem to more uniformly range through the habitat.

Unfortunately, none of the habitat variables we assessed were very effective in

explaining the variations in drill presence along the transects. The strongest associations

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we found were with forest type, which is a factor of the high proportion of points with drill presence in the Caldera, and river proximity. However, diagnostic plots indicate a

poor fit, and high residual deviance denotes a weak explanatory value of the model. The

most likely explanation for our inability to identify which variables were most associated

with microhabitat use is related to differences in the distribution of encounters along each trail and foraging patterns we previously discussed.

Unlike at either of the lowland forest sites, drills in the Caldera are rarely encountered at the same location on consecutive days. In addition, the primary food items

of drills in the Caldera are nearly ubiquitously found throughout the habitat, indicating that foraging patterns may be more randomly dispersed. Conversely, drills in lowland forests were often observed visiting the same patch of fruiting trees at least once a day, over the course of several days or weeks. Along Badja trail, the patchy distribution of encounters and high frequency of presence at a few points indicates that drills may be repeatedly using the same, or similar, routes to access fruiting trees. Primates have been shown to possess exceptional spatial and temporal memory, often enabling them to travel the shortest, linear routes to distant food sources (Altmann 2009). If drills in lowland forests utilize such an optimal foraging pathway, their presence at a given point along

Badja trail might simply be a consequence of their path selection, and be more related to the linear distance between feeding trees than the habitat variables at any point.

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Conclusions

Bioko Island drills were found to have low group sizes and rates of polyspecific

encounters, a high occurrence of single-male/single-female groups compared to drills and

mandrills on the mainland. The similarity between these findings and those of other primates, as well as their cryptic defensive behaviors and the high abundance of sympatric frugivorous primates on Bioko Island suggest that these differences are related to three factors: 1) high human predation from shotgun hunting, 2) an absence of non- human predators, and 3) high interspecific competition.

These findings have important implications to the conservation of drills on Bioko

Island. The decreased ratio of adult males to females suggests that the overall reproductive output of each group is relatively low compared to the mainland subspecies.

In semi-captive conditions at the Drill Rehabilitation and Breeding Center in Calabar and

Afi Mountain (DRBC), Nigeria, females birthed one infant at a time, at a mean interbirth

interval of 473 days (0.77 births/year) (Wood 2007). Based on our maximum mean group

size estimate (5.3 individuals), the average number of female Bioko Island drills per

group is 1.2. If we assume that the birthrate reported in the DRBC holds true for wild

drills on Bioko Island, the average group produces 0.91 infants per year. Given that this

birth rate is roughly 4.6 times the contemporary rate that drill carcasses are being sold in

the bushmeat market, it is very likely that unless action is quickly taken, drills will soon

be extirpated from Bioko Island.

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Table 4.1: Summary information for drill group sizes and compositions between the Caldera, Moaba Playa, and Moraka Playa and survey methods, including the number of groups encountered and the mean number of individuals in each group. The average proportion and maximum number of individuals of each member class. Statistical significance values (Kruskal- Wallis test) are provided for any significant comparisons. Encounters Group Male Female Sub-Male Juvenile Infant Unknown Survey Type (N) Size mean (+SEM) max 2.0 (0.2) 29.7 (3.3) 31.4 (3.1) 13.7 (2.4) 8.1 (1.8) 1.6 (0.7) 15.4 (2.9) Total Transects 143 18 2 7 4 6 3 2

Reliable 4.2 (0.5) 32.3 (5.4) 33.5 (3.4) 15.2 (3.4) 12.0 (2.9) 4.3 (1.8) 2.8 (1.6) 34 Transects 18 2 7 4 6 3 2

Blinds 3.4 (0.3) 20.4 (3.5) 18.7 (2.1) 34.1 (4.1) 11.8 (2.1) 7.5 (1.3) 7.6 (2.0) 102 (Moaba Playa) 20 2 4 4 7 4 8

Reliable 3.6 (0.3) 23.4 (3.0) 22.4 (1.9) 29.3 (3.3) 11.7 (1.7) 6.7 (1.1) 6.4 (1.6) 136 Encounters 20 2 7 4 7 4 8 Statistical Comparisons Site Differences ns ns ns ns ns ns ns

H=5.9, H=13.6, H=12.8, ns ns ns ns Transect versus Blind p = 0.02 p < 0.001 p < 0.001

Transect H=31.5, H=5.7, H=6.3, H=6.0, ns ns ns (Total versus Reliable) p < 0.001 p = 0.02 p = 0.01 p = 0.01 111

Table 4.2: The proportion (%) of polyspecific associations recorded during encounters with Bioko Island drills (N = 143) and drills in Korup N.P., Cameroon (N = 44), as reported by Astaras et al. (2011). Proportion of Encounters (%) Species Common Name Bioko drills Mainland drills Cercopithecus erythrotis Red-eared monkey 3.5 30 Cercopithecus pogonias Crowned monkey 3.5 25 Cercopithecus nictitans Putty-nosed monkey 0 39 Allocrocebus preussi Preuss’s monkey 1.4 - Procolobus pennantii Red colobus 3.5 - Colobus satanas Black colobus 4.9 - Cercocebus torquatus Red-capped mangabey - 25 Piliocolobus preussi Preuss’s red colobus - 9 Cercopithecus mona Mona monkey - 14 Polyspecific Encounters 14 50 112

Appendix 4.1: Group size information on drills and other Papionin primates, including mandrills, mangabeys (Cercocebus), baboons (Papio), and geladas (Therapithecus). Group Sizes Study Site Species Mean Range Astaras et al. 2008 Korup N.P., Cameroon Mandrillus leucophaeus 52.3 25-77 Gartlan 1970 Bakundu F.R., Cameroon Mandrillus leucophaeus 63.5 14-179 Wild et al., 2005 Bakossiland, Cameroon Mandrillus leucophaeus 93.1 5-400 Schaaf et al., 1990 Bioko Island Mandrillus leucophaeus ND 1-7 Gonzalez-Kirchner & de la Maza 1996 Bioko Island Mandrillus leucophaeus ND 2-20 Other Papionins Tutin et al. 1997 Lopé, Gabon Mandrillus sphinx 150 No data Abernethy et al. 2002 Lopé, Gabon Mandrillus sphinx 620 338-845 Matthews & Matthews, 2002 Campo, Cameroon Mandrillus sphinx ND 15->100 Mbora et al., 2009 Tana River, Kenya Cercocebus galeritus 32.2 25.8-40.3* Mitani, 1989 Campo, Cameroon Cercocebus torquatus 21.1 14-38 Devreese, 2011 Bai Hokou, CAF Cercocebus agilis 134.2 130-136 Bronikowski & Altmann, 1996 Amboseli, Kenya Papio cynocephalus 59.7 47-78 Freeland, 1979 Kibale, Uganda Papio anubis 34.3 26-46 Iwamoto & Dunbar, 1983 Ethiopoa Theropithecus gelada 47.2 14.9-51.1* * Denotes the range of average group sizes, when absolute range information was not available 113

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CHAPTER 5: STABLE ISOTOPE ECOLOGY

Introduction

Throughout ecological and evolutionary science, competition has been thought of

as one of the primary mechanisms driving behavioral and morphological adaptations, and

critical to the maintenance of phenotypic and genetic variation in species. Darwin

highlighted its importance, stating,

“Natural selection, also, leads to divergence of character; for more living beings can be supported on the same area the more they diverge in structure, habits, and constitution, of which we see proof by looking at the inhabitants of any small spot or at naturalized productions. Therefore during the modification of the descendants of any one species, and during the incessant struggle of all species to increase in numbers, the more diversified these descendants become, the better will be their chance of succeeding in the battle of life. Thus the small differences distinguishing varieties of the same species, will steadily tend to increase till they come to equal the greater differences between species of the same genus, or even of distinct genera” (Darwin, 1859, p. 169).

Subsequent studies on the ecology and behavior of the Galapagos ground finches

have continued to provide evidence to support and expand Darwin’s early hypothesis

(Lack 1945; Abbott et al. 1977; Grant & Grant 1982, 2006), as have numerous field and

experimental studies on other avian and non-avian taxa (Brown & Wilson 1956; Schluter

& McPhail 1992; Schluter 2000) (Reviewed by: (Dayan & Simberloff 2005)).

Despite considerable debate throughout the ecological literature on the role of

competition in evolution and community dynamics (Jr et al. 1979; Grant & Abbott 1980;

Connell 1983; Roughgarden 1983), there has been a general consensus on the importance

of variations in niche width and overlap to the coexistence of species competing for

limited resources (Pielou 1972). Behavioral or morphological differences that enable

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species to partition shared resources or utilize alternatives such that they mitigate the

effects of competition are likely to benefit fitness and promote the maintenance of species

richness (Svanbäck & Bolnick 2007). Thus, competition and niche theory have often been

employed to address a variety of questions involving the species abundance and richness of communities (MacArthur et al. 1972), speciation and phenotypic variations (Bolnick et

al. 2003; Berg & Ellers 2010), and interspecific ecological differences (Gartlan &

Struhsaker 1972; Schreier et al. 2009). Niche theory has often been used to describe

species generally, with little regard to intraspecific variations (Bolnick et al. 2003).

However, theoretical models (van Valen 1965) and experimental evidence (Bolnick 2001;

Svanbäck & Bolnick 2007) have provided support for the potentially strong influence of

intraspecific competition on driving individual specialization and increasing the niche

breadth of a population. Therefore, investigations of the impact of competition on community and niche dynamics should incorporate both intra– and interspecific variations.

One of the primary tenets of island biogeography is that there is a positive

correlation between the area of an island and the number of species present (MacArthur &

Wilson 1967; Hubbell 2001). This low species richness on islands may decrease the

number of species competing for similar resources, compared to mainland communities

(MacArthur et al. 1972). As a result of this competitive release, insular populations are

often found in higher abundances and with shifted or expanded niches compared to

conspecifics living in adjacent species-rich ranges (Diamond 1970a, 1970b). The

community dynamics related to this species-area relationship has made islands a

particularly important and frequently used system to study niche variation and community

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structures. However, the effects can vary considerably between species and systems, and

likely involve a complex set of ecological interactions (MacArthur et al. 1972). In a study

of “forest islands” in Southern Dakota, Martin (1981) reported species belonging to species-rich ecological guilds were present in fewer islands compared to species in guilds with fewer members. He argued that the limited species richness and abundance in these fragments led to community compositions that were structured such that there were fewer species belonging to the same guild. Ultimately, the composition of an ecological community involves the variations in the traits and competitive interactions between species, and local environmental conditions (Martin 1981).

Bioko Island, Equatorial Guinea is a continental shelf island approximately 32 km off the coast of Cameroon in the Gulf of Guinea. The island separated from the mainland approximately 10,000 to 11,000 years ago during the glacial recession of the late

Pleistocene Epoch (Sayer et al. 1992). Currently, eleven species of primates (including seven cercopithecine monkeys and four galagos) live within the forests of Bioko Island; however as 27 species live within the proximate mainland forest habitat of Cameroon, the current richness is likely a small subset of the level prior to their separation.

Based on the species-area curve equation, Cowlishaw (1999) calculated the expected number of primates within the 2017 km2 area of Bioko Island to include only

five species, a reduction of 22 following its insularization. A total of nine primate species

were known to exist on the island at the time of this study, leading Cowlishaw to suggest

that 67% of the original species had already been extirpated, and that an additional 15%

were likely to be lost. Following the publication of this study, two additional species have

been confirmed on the island, including Thomas’s dwarf galago (Galagoides thomasi)

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and the Bioko needle-clawed galago (Euoticus pallidus pallidus) (Grubb et al. 2003).

Given that there are now 11 primate species recognized on Bioko Island, we should

expect an additional 22.2% reduction of its pre-separation richness, a loss of more than

half the species currently present (6/11).

Within the Gran Caldera and Southern Highlands Scientific Reserve, a largely

unprotected reserve comprising the southern third of Bioko Island, the relative density of

anthropoid primates is among the highest reported in the world. Butynski and Koster

(1994) estimated total densities at 2.02 encounters/km-1, and although more recent work

has documented a decline in abundances in this area, they remain relatively high (Cronin

2013). Given the relatively high species density and richness of monkeys on Bioko, the

potential for competition and niche overlap on the island may be high. The cercopithecine

monkey species present on Bioko fall within three dietary guilds; (1) granivorous-

folivores– the black colobus (Colobus satanas) and Pennant’s red colobus (Procolobus

pennantii), (2) arboreal frugivorous-omnivores (guenons)– the Preuss’s monkey

(Allochrocebus preussi), putty-nosed monkey (Cercopithecus nictitans), crowned monkey

(C. pogonias), and the red-eared monkey (C. erythrotis), and (3) a single terrestrial

frugivorous-omnivore– the drill (Mandrillus leucophaeus) (Fa & Purvis 1997). Very little

research has been performed on the ecology or behavior of the primates of Bioko Island,

and much of the available information is either admittedly preliminary or anecdotal in

nature (Butynski & Koster 1989; Schaaf et al. 1990; Mate & Colell 1995; Gonzalez-

Kirchner 1996; Gonzalez-Kirchner & de la Maza 1996). However, reports from mainland

populations (Struhsaker 1969, 1975; Gautier-Hion et al. 1997; Chapman & Chapman

2000a) and observations made during this study indicate that although the primary dietary

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components of these three guilds are different, there is considerable overlap between the

food types and species consumed.

Recent research into the ecology and behavior of the Bioko Island drill subspecies

(M. l. poensis) has identified several dietary and behavioral characteristics that differ

significantly from what has been reported on the mainland subspecies (M. l. leucophaeus)

(Chapters 2 & 4 of this dissertation). Numerous differences in the diet were found, most

notably was that the proportion of the dry weight of fecal samples collected within

montane forest habitat on Bioko Island were 45% higher in non-fruit fiber and 61% lower

in fruit than the average reported on the mainland (Astaras 2009). The population

differences in foraging groups size and composition were even more dramatic, as even the

highest estimated mean number of individuals per group on Bioko (5.3) was only 7.6% of

the average size of groups reported on the mainland (69.6) (Astaras et al. 2008, 2011;

Morgan et al. 2013). In addition, the average ratios of males to females and males to all

group members on Bioko (1:1 and 1:4) were considerably lower than mainland drills (1:8

and 1:20), and polyspecific associations occurred in 36% less encounters on the island

(Astaras 2009).

One potential explanation for these dietary and behavioral differences between the

insular and mainland drill subspecies is the relatively high abundance and richness of fruit

competitors on the island. Each of the other six monkey species are potential competitors

for fruit resources, and numerous non-primate frugivores exist on the island, including the

blue duiker (Philantomba monticola) and Ogilby’s duiker (Cephalophus ogilbyi). These forest antelopes are often observed consuming the same fruit and vegetative foods as the primate species, especially fruits of Ficus spp. P. monticola, in particular, has been

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described as one of the most frugivorous species of ruminants (Kendrick et al. 2009).

Stomach content analysis of 16 individuals in Gabon found fruit and seeds to compose

71% of the average total weight of the food ingested (Dubost 1984). The high potential

for competition may explain the dietary variations recorded in Bioko Island drills, driving

them to consume alternative diets, and resulting in changes in the group size.

Unfortunately, poor visibility in the forests of Bioko Island, high hunting pressure,

and a lack of habituated groups currently make detailed comparative observational studies

an unrealistic endeavor. Comparative studies using fecal remains are also unlikely to be

relatively informative, as both the colobus and guenon species are primarily arboreal and

flee from human contact, making fecal sample collection much more difficult than with

drills.

In addition, both the colobus and duikers have specialized digestive systems that

include a large, sacculated forestomach, where bacterial fermentation enables the

digestion of the cellulose of plant tissues (Davies & Oates 1994). This differential

digestion makes comparisons of fecal samples collected from the colobus or duikers with

those from other monkey species unreliable.

Increasingly, stable isotope analysis has been used to address questions of niche

differentiation and overlap, and investigate inter- and intraspecific competition. Stable

isotope ecology rests on the basic principle that you are what you eat, and therefore

ecological information can be ascertained from the analysis of body tissues. More

specifically, the ratios of heavy to light isotopes of an element leave a signature within

consumer tissues proportional to those of its dietary items. Measurable differences in the

signatures of food items are identifiable in consumer tissues, and thus provide a record of

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their diet (Jardine et al., 2006). This technique provides a quantitative comparative metric

to investigate the ecological niches of species that would otherwise be limited by most

traditional methods, particularly with elusive or rare species, for which the number of

observations or capture evens would be minimal (Hilderbrand 1996; McFadden &

Sambrotto 2006). Although stable isotope analyses have only been recently incorporated

into the toolbox of ecologists, a large number of studies have successfully addressed a

broad range of questions, showing that this tool can be highly accurate and informative

(Crawford et al., 2008).

The most commonly used stable isotopes in ecological studies are carbon13 and nitrogen15, denoted as the per mil fraction of heavy to light isotopes (δ13C‰ and δ15N‰).

The δ13C values of plant tissues, and consequently their consumers, vary consistently in relation to the photosynthetic pathway used. The differential fractionation of atmospheric

13 carbon between C3 and C4 plants results in non-overlapping ranges of δ C values

between -34‰ to -22‰ (mean=-27.1+2.0) for C3 plants and -15‰ to -9‰

13 (mean= -13.1+1.2) for C4 plants (Farquhar et al. 1989). Within these ranges, δ C has

been shown to decrease due to local environmental factors, including lower light levels,

higher moisture, lower temperature, and higher partial pressures of atmospheric gasses

(Heaton 1999; Sandberg et al. 2012). These factors have been shown to vary carbon

isotope values of plants along gradients in vertical forest strata and altitude. Plants on the forest floor can be depleted as much as 2‰ to 5‰, compared to those in the upper canopy

(referred to as the “canopy effect”) (Broadmeadow et al. 1992), and increasing elevation has also been shown to correlate to carbon enrichment by an average range of +1.0-1.6‰ per km gained (Körner et al. 1988, 1991; Hultine & Marshall 2000).

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Variations in δ15N of plant tissues is less understood than δ13C, however they are

most strongly influenced by the local climatic conditions, with mean annual temperature

and rainfall correlated to δ15N depletion in plant tissues (Amundson et al. 2003).

Leguminous plants are typically lower in their δ15N values than other plants, although this

is not true for all species (Delwiche et al. 1979; Virginia & Delwiche 1982). However,

because a relatively consistent nitrogen discrimination of +3‰ to +4‰ between trophic

levels has been recorded in an array of species, δ15N is most often used in ecological

studies to distinguish and compare the trophic position of consumers (e.g.,(Schoeninger et

al. 1997; McFadden & Sambrotto 2006; Rodríguez & Gerardo Herrera 2013; Pearson et

al. 2013)).

Here we used stable isotope analysis of carbon and nitrogen to compare the niches

of representative species from each of the three guilds present in the Bioko Island

cercopithecine community (M. leucophaeus, C. satanas, and C. erythrotis) and the blue

duiker (P. monticola). Analysis was performed on hair samples collected from dead

animals being sold at the bushmeat market, which were hunted at locations throughout

Bioko Island. We used known dietary information from fecal analysis of the drill to

interpret the stable isotope values, and compare the results between taxonomic,

demographic, spatial and temporal scales. Our objectives were to compare the niche

breadth and overlap between species, sex classes, and seasons, and characterize the

species specific dietary differences. We used this information to assess the potential

competition between species and identify strategies used to mitigate its effects. We also

used the known dietary differences of the drill between montane and lowland forests to

determine if nitrogen or carbon isotopes can be used to identify where individuals were

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hunted.

Methods

Ethical Note

Samples were obtained from the carcasses of dead individuals being sold to local

consumers during the sale process. The collection of samples from illegally hunted

endangered species raises ethical concerns related to the stimulation or support of such

markets. The methods used in this research were designed and executed to ensure that

they did not promote hunting or sale of bushmeat. No money, gifts, or other incentives

were given to the individuals selling, preparing, or purchasing the carcasses. Instead, sample collections were performed as part of a partnership with the National University of Equatorial Guinea (UNGE), School of Environmental Studies. The faculty and students of UNGE are highly regarded among local people on Bioko Island, which enabled us to gain access to the carcasses for sampling purposes. Partners from UNGE were trained on the methods during the first four months of the sampling period, and

continued on their own until sampling was completed.

Sample Collection and Preparation

Hair samples were collected from January 2010 to May 2011 at Semu Bushmeat

market in the capital city of Malabo, which is located on the northern coast of Bioko

Island. Roughly 50 hairs were clipped from the rump of each individual, just before the base of the tail, as close to the skin as possible using fine tipped dissection scissors

(Schoeninger, 1998; Dammhahn and Kappeler, 2009). Between each collection, scissors

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were cleaned with ethanol and gloves were changed. Upon removal, samples were

manually cleaned of any debris, placed in sample collection bags, and stored in sealed

bags containing silica gel. Hair samples were rinsed in a bath of 70% Ethanol and dried

overnight for 24 hours at <60°C. Samples (~0.5 mg) of whole hairs of an approximately

equal length were transferred into aluminum capsules for isotope analysis. Hair is a

metabolically inert tissue with a high isotopic turnover rate that can record several months

of stable isotope data (Michener & Lajtha 2007; Dammhahn & Kappeler 2010). Samples

were analyzed from two collection periods, those collected from the market from January

to May were considered dry season samples, and those from July through November were

wet season samples.

A total 303 hair samples were collected from the carcasses of seven monkey

species and two duikers on Bioko Island. The species representing each guild in the

analysis were selected based on the sample sizes collected, those with the largest N were

used (C. erythrotis (98), M. leucophaeus (54), P. monticola (48), C. satanas (26)). We

sought to analyze five individuals of each sex class, for both seasons. However, we were

unable to collect P. monticola samples during the wet season, and only sampled one

female C. satanas during the wet season, resulting in variations in the number analyzed

between species and sexes.

As isotope signatures vary due to local environmental conditions, the baseline

signatures of primary producers are needed to enable comparisons with studies performed

in other locations. Such data are also necessary for the isotopic mixing model used in this

study to assess the proportional contribution of food items. To develop this baseline, the

stems, leaves, fruits, and roots of a variety of plants and whole insects known or

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suspected to be consumed by the focal consumers were collected opportunistically in the

field. Samples were only collected during the dry season, from January to March 2012,

and were taken from a variety of elevations to test for altitudinal variations in isotope

values. Plant and invertebrate samples were dried in the field (<50°C) in aluminum foil

boats and transferred to sealed bags which were stored in containers with silica gel. Prior

to analysis, the samples were homogenized using a ball mill and ~1.5 mg of the

homogenate loaded into aluminum capsules.

Mass spectrometer analyses were carried out by the Analytical Chemistry

Laboratory at the Odum School of Ecology, University of Georgia. Ratios of 13C/12C

and 15N/14N and the total carbon and nitrogen content of the samples were analyzed

using the Carlo Erba Elemental Analyzer (NA 1500) and Finnigan Delta C. continuous

flow isotope ratio mass spectrometer. All samples were analyzed in duplicate. Isotope

ratios were reported in the standard per mill (‰) delta notation (δ) relative to the

international standards PDB (carbon) and Air (nitrogen), calculated as δX‰ =

(Rsample/Rstandard - 1) · 1,000; where Rsample and Rstandard are the molar ratios of

heavy to light isotopes for the sample and standard respectively (Martinez del Rio et al.,

2009).

As food items are digested and the nitrogen and carbon are incorporated into tissues, the isotopic signatures are changed due to physiological and chemical processes

(Michener & Lajtha 2007). To correct for this fractionation, trophic enrichment factors

(TEF) were utilized: (TEF=δ tissue - δ diet). TEF’s have been found to vary considerably

between species. However controlled experimental studies with several mammalian

herbivores have shown hair to be 3‰ higher for carbon and 3.6‰ for nitrogen, compared

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to the dietary items (Sponheimer et al. 2003a, 2003b). Recent studies have reported

similar TEF values in wild primates (Oelze et al. 2011; Nakashita et al. 2013), and as a result we have applied them to our analyses.

Data Analysis

All comparisons were made using the mean δ13C, δ15N, Total C% and Total N% of the sample duplicates for each individual. Differences in the δ13C and δ15N between

the hair samples of different species, sex classes, and seasons were compared

independently using one-way analysis of variance tests (ANOVA). Seasonal comparisons

were not possible with P. monticola, as we were unable to collect any hair samples

during the wet season. Subsequent multiple comparisons of the components were made

using Tukey’s honest significance test (Tukey HSD) in order to identify the significant

comparisons. Differences in the stable carbon and nitrogen isotope values, as well as the

total contribution of carbon (%C) and nitrogen (%N) of the plant and invertebrate tissues

were compared either through one-way ANOVAs and Tukey HSD tests, or Kruskal-

Wallis tests when data were found to be non-parametric. Subsequent multiple

comparisons were made using kruskalmc from R package pgirmess (Giraudoux 20012).

Linear regressions were used to determine the response of the isotope signatures of plant

and invertebrate tissues to increasing altitude.

Differences in the isotopic niche space (also referred to as δ space) occupied by each species were compared using methods recently adapted from the ecomorphology literature to isotope ecology, whereby niche characteristics are quantified from the two-

dimensional bi-plots of the δ13C-δ15N space (Hoeinghaus & Zeug 2008). To compare the

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total trophic diversity, we generated minimum convex hulls created by the samples from

individuals of each species and calculated the total area (TA) covered each hull

(Hoeinghaus & Zeug 2008). As convex hulls are generated from the individuals at the

extreme value ranges, the size and shape of hulls will be dramatically altered by one or

more outliers in the population. To assess the distribution of individuals within each hull,

the mean nearest neighbor distance (NND) between each individual within a species was

calculated. A relatively larger TA represents a species with a broader overall trophic

diversity, whereas a low mean NND indicates a clustered distribution within the space

(Hoeinghaus & Zeug 2008). Individual nearest neighbor values were compared between

species using a one-way ANOVA and subsequent comparisons made through Tukey

HSD. Kernel density estimates were plotted to visualize the two-dimensional (2D)

distribution of individuals within the niche space and the separate one-dimensional

frequency distributions (histograms) of δ13C and δ15N values. The distributions were

plotted via the R package ggplot2 (functions stat_density2d and stat_density)(Wickham

2009).

The relative proportions of food sources to the diets of each species were determined using the package ‘Stable Isotope Analysis in R’ (SIAR) (Parnell et al. 2008).

SIAR uses a Bayesian framework to develop probability distributions of multiple food sources in the diet of consumers. SIAR is the most commonly used isotopic mixing model software because it enables users to incorporate trophic enrichment factors between the source and consumer isotope values, as well as variations in the elemental concentrations of the tissues (Parnell et al. 2010). As previously discussed, we set the trophic discrimination factor at 3‰ for carbon and 3.6‰ for nitrogen (Sponheimer et al.

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2003a, 2003b), and used the mean %C and %N for the elemental concentration values of each source. Only hair samples collected from the dry season were used, as the lack of source materials from wet season may skew the results of the mixing model. SIAR generates diagnostic plots which provide the coefficients of correlation in the posterior distributions of the covariates (sources) for each consumer which we will use to determine the model’s strength (Parnell et al. 2010).

All analyses were performed using R statistical software v 2.15.0 (R Core

Development Team 2012). Significance tests were two-tailed with the significance level set at 0.05.

Results

Food Sources

The mean stable isotope values of the food source samples varied between -

34.18‰ to -11.4‰ for carbon and 0.54‰ to 5.32‰ for nitrogen (Table 5.1).

Comparisons between the food items revealed several significant differences in their isotope values (Table 5.2), most notably were the differences between C4 grasses, invertebrates, and the combined tissues of C3 plants (F(5,80)=117.37, p< 0.001) (Figure

5.1). The mean δ13C value of C4 grasses (=-11.36, SD=0.46) was significantly higher than all other source types (Tukey HSD, p<0.001 for all comparisons). Average δ13C

values were lowest in the tissues of the herbaceous groundcovers (=-34.18, SD=0.71), significantly lower than Aframomum spp., invertebrates, and trees (Tukey HSD, p<0.001

for each).

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Invertebrate samples were highly enriched in δ15N (=5.32, SD=1.0), ranging

between 3.94‰ and 6.64‰, and were significantly higher than all other food types

except the herbaceous groundcovers (=3.56, SD=1.5) (F5,80=14.12, p< 0.001; Tukey

HSD, p<0.001 for all comparisons). The δ15N values in the groundcover plants were significantly higher than Aframomum spp. (Tukey HSD, p=0.002) and the tissues of tree species (Tukey HSD, p=0.01).

The proportion of carbon and nitrogen to the total sample mass was also significantly different between food types (%C: F5,80=7.58, p< 0.001; %N: F5,80=374.3,

p< 0.001) (Figure 5.1). Invertebrate tissues contained the highest proportion of carbon by

mass; significantly higher than C3 plants (Tukey’s HSD, p=0.03), C4 grasses (Tukey’s

HSD, p=0.02), groundcovers (Tukey’s HSD, p<0.001), Aframomum spp. (Tukey’s HSD,

p=0.004), and roots (Tukey’s HSD, p=0.002). The nitrogen content in invertebrate tissues

was also significantly higher than all other food parts (Tukey’s HSD, p<0.001), over

seven times as much as the combined average of plant samples (Table 5.1). Nitrogen was

lowest in Aframomum spp., only making up an average 0.7% of the sample mass. This

value was significantly lower than C4 grasses (Tukey’s HSD, p=0.02), groundcovers

(Tukey’s HSD, p<0.001), and trees (Tukey’s HSD, p=0.001).

Comparison of the δ13C values of the terrestrial herbaceous plants collected

(Aframomum spp., Paracostus englerianus, and Dracaena phrynioides) were positively

2 correlated with elevation (R =0.52, F1,14=17.28, p<0.001), increasing by 4.19‰ for every kilometer gained (Figure 5.2). This relationship was not found in the invertebrate tissues,

or with the δ15N values of any food types.

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Consumer Isotopic Variations

The stable isotope values of the individual consumer hair samples varied between

-7.6‰ to -22.7‰ for carbon and 2.6‰ to 8.6‰ for nitrogen (Table 5.3). Average δ13C values were highest in C. erythrotis (-24.6‰) and lowest in P. monticola (-25.8‰), and

δ15N was highest in P. monticola (6.6‰) and lowest in C. satanas (4.6‰) (Figure 5.3).

Significant differences existed between species in the values of both isotopes (δ13C:

15 13 F3,69=3.82, p=0.01; δ N: F3,69=12.02, p< 0.001). Pairwise comparisons show the δ C values of P. monticola to be significantly lower than C. erythrotis (p=0.008) and M. leucophaeus (p=0.05), and the δ15N values of P. monticola and C. erythrotis significantly

higher than M. leucophaeus (P. monticola: p=0.002; C. erythrotis: p=0.007) and C.

satanas (both: p<0.001).

Bi-plots of the isotope niche space (δ13C-δ15N) encompassed by each species

show considerable interspecific overlap in the δ-space occupied (Figure 5.4). The

minimum convex hulls of C. satanas occupied the largest total niche area (TA=15.95),

followed by C. erythrotis (TA=9.18), M. leucophaeus, (TA=6.3), and P. monticola

(TA=4.68). Mean nearest neighbor distances were highest in C. satanas (1.96,

SEM=0.08) and P. monticola (1.94, SEM=0.15), and significant differences existed between species (F(3,671)=13.79, p<0.001). Multiple comparisons show the distances of M.

leucophaeus (1.34, SEM=0.06) to be significantly lower than all other species (Tukey

HSD, p<0.001 for all comparisons), and the values of C. erythrotis (1.67, SEM=0.07)

lower than C. satanas (p<0.02). Plots of the kernel density contours show C. satanas and

M. leucophaeus to have the smallest 2D density probabilities, and that the relatively large

convex hull of C. satanas is a result of two males and two females that had δ-values

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distant from the otherwise clustered population (Figure 5.5). The 2D density of P.

monticola was more evenly distributed throughout a large area of the niche space, with

no individuals outside of the probability contours.

No significant differences were found in the mean δ13C or δ15N values between the sex classes or seasons of any species. However, the total niche space area occupied

and the mean nearest neighbor distances were both an average 1.4 times higher in males

than females (Figure 5.4). Nearest neighbor distances were significantly higher in the

males of C. erythrotis (F(1,108)=10.11, p=0.002), C. satanas (F(1,18)=5.34, p=0.02), and M.

leucophaeus (F(1,108)=12.07, p<0.001). Lack of significant differences in the sex classes of

P. monticola are likely due to the low sample size (N: ♂=5, ♀=5), rather than niche

similarity. Density histograms for the frequency of both isotopes were, in general,

smoother for females (Figure 5.5). This was especially true for nitrogen, with males

displaying considerable variability over a wider range of δ15N values. Total niche area

declined from the wet season to the dry season in C. erythrotis and M. leucophaeus, and increased in C. satanas (Table 5.4). Nearest neighbor distances were significantly higher

in drill samples from the wet season (F(1,98)=8.71, p=0.004), however no other differences were significant.

Dietary Source Proportions

Based on the statistical differences in the isotope values of the food items and dietary information from fecal sample analysis and observations, sources were aggregated into five groups (C4 grasses, Aframomum spp., groundcovers, other C3 plants, and invertebrates) to use in the mixing models. The isotope values of the consumer

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species fit within the mixing polygon of the source errors, and primarily fell in a band

between the C3 plants and invertebrates (Figure 5.6). Correlations between the posterior

distributions of the food items were high between the predicted contributions of several

food sources to the diets of the consumers. The majority of the strong negative

correlations (coefficients >0.6) were particularly those between the groundcovers,

Aframomum sp., and the other C3 plants, indicating that the model may have difficulties

discriminating between these sources. Positive correlations, which indicate that the model

necessarily required both sources to balance each other out and generate a solution (Inger

et al. 2006), existed only between C4 grasses and groundcovers.

Even with the suspect fit of the model, several patterns are apparent in the

predicted proportions of each source to the consumer diets (Figure 5.7). First, C4 grasses

are likely non-existent in the diets of any species, despite the SIAR boxplots showing the

predicted mean contribution of roughly 15-20% to each diet. This is likely an artifact of

the strong positive correlations between C4 grasses and the groundcovers, as none of the

δ13C values of any of the consumers are close to the range of C4 consumers. Second, C3

plants, including Aframomum spp. and the groundcovers, make up the majority of all

species’ diets. Third, both groundcover and invertebrate consumption is highest in P.

monticola (Figure 5.7). Attempts to modify the model by removing or combining sources

to increase the fit were unsuccessful. This indicates that either we failed to include certain

important food sources in our analysis, or that the high variation in isotope values, particularly of the C3 plants, may limit the model’s ability to discriminate between the sources.

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Discussion

The results of the stable isotope analysis of basal resources on Bioko Island were

similar to the values found in other studies. Mean δ13C values of C3 (-30.5, SD +2.6) and

C4 (-11.4, SD +0.5) plants fell within the expected range (Farquhar et al. 1989). Carbon

isotope values in groundcover species were 4.6 ‰ less than tree species, and fruits were

the most δ13C enriched vegetative tissues in our study, all of which is consistent with the

expectations of the canopy effect. Although Aframomum spp. are also low-growing

herbaceous plants, they primarily grow in large patches with open canopies and are not

subject to the factors of the canopy effect, explaining their relatively enriched δ13C values

compared to the groundcovers.

The rate of increase in δ13C values of herbaceous plants with altitude is dramatically higher than previous studies, which reported mean rates of approximately

+1.0-1.6‰ per km, ranging up to +2.68‰ (Körner et al. 1988, 1991; Hultine & Marshall

2000). Altitudinal enrichment of δ13C values is attributed to a combined effect of

decreased temperature and the partial pressure of oxygen, which result in increased

discrimination of δ13C during photosynthesis (Körner et al. 1991). Altitudinal gradients in

the partial pressure of atmospheric gasses and atmospheric temperature are relatively

consistent across the world (Körner 2007), and there is no evidence to suggest that either of these factors vary more on Bioko Island, than in any other study regions. We currently have no explanation for this elevated rate of change, however as these results were derived from a limited number of species (3) and samples (16), it is possible that other local environmental variations may be at work.

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Similar to previous reports, mean δ15N values of different plant species and

tissues on Bioko Island were highly variable (Sandberg et al. 2012); the standard

deviation of the combined plant values (+0.81) was equal to 62% of the mean (1.30).

However, compared to the values reported of plants in other sub-Saharan tropical forests,

the basal level of nitrogen isotopes on Bioko Island were depleted by an average of 4.0

‰ (Cerling et al. 2004; Oelze et al. 2011; Blumenthal et al. 2012). While numerous

environmental factors, including temperature, topography, and the age and sediment type of soils have been shown to influence geographic variations in the nitrogen isotopes of

plants, a strong negative correlation exists between δ15N values and mean annual

precipitation at a location (Amundson et al. 2003). Annual precipitation on Bioko Island

is considerably higher than proximate mainland forests, and the duration of the single wet

season is longer due to oceanic influences (Adams 1957). As increased mean annual

precipitation has consistently been shown to decrease plant δ15N values, the depletion of

baseline nitrogen isotope signatures relative to mainland sites is not surprising

(Amundson et al. 2003).

Consumer Values

Differences in the carbon isotope values between P. monticola and primate

species provide some evidence that the canopy effect may be recorded in consumer

tissues on Bioko. δ13C ‰ was lowest in P. monticola, significantly less than the species

with the highest mean value, C. erythrotis, by 1.2 ‰ on average. As an ungulate, P.

monticola was the only species studied to forage solely on the forest floor and was regularly observed consuming the stems and leaves of terrestrial herbaceous vegetation,

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resulting in δ13C depletion. However, the difference in δ13C ‰ is less than that found

between the herbaceous groundcover and tree species, which is likely a result of the large

contribution of fallen fruits in the diets of P. monticola, as has been found in other studies

(Oelze et al. 2011). In a community wide isotope study of mammals in the Ituri Forest,

Democratic Republic of Congo, no significant difference were found in carbon isotope values between seven duiker species (including P. monticola) and any other species

(including six primates), with the exception of other ungulate species which primarily consumed the leaves of subcanopy vegetation (Cerling et al. 2004).

The lack of significant differences between the carbon isotope values of the primate species are more difficult to explain. Published studies of the vertical distributions of C. satanas, C. erythrotis, and M. leucophaeus suggest that they differ dramatically in their use of the forest strata. On Bioko Island, C. satanas were observed in the middle, upper, and emergent canopy (>20 m from the ground) during roughly 75% of encounters, whereas C. erythrotis groups were in the lower canopy or forest understory

(<20 m from the ground) in 80% of encounters (Gonzalez-Kirchner 1996, 1997). M.

leucophaeus are the most terrestrial monkey species within their range, and were reported to be on the ground in 73.3% of observations on Bioko Island (Gonzalez-Kirchner & de la Maza 1996), and within three meters of the ground in 63% of encounters in Korup

National Park, Cameroon (Astaras et al. 2011). These values are also consistent with unpublished observations on Bioko Island (G. W. Hearn, pers. obs.). δ13C values do not

reflect these differences, instead they suggest C. erythrotis are highest, followed by M.

leucophaeus, and C. satanas. However, dietary variations of these species suggests that

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the depleted δ13C values of C. satanas may be a result of differences in food items

consumed, despite originating in higher forest strata.

Compared to the other primates on Bioko Island, the specialized digestive systems

of Colobus monkeys enables them to better digest and extract nutrients from leaves.

Studies of C. satanas have reported leaves to make up roughly 20 to 40% of all food

items, whereas fruit consumption was infrequent (Gautier-Hion et al. 1997; Tutin et al.

1997). This is considerably different from the frugivorous-omnivore diets of the guenons

(C. erythrotis) or M. leucophaeus. Given that we found the δ13C values of leaves to be significantly lower than fruits, the increased consumption of leaves by C. satanas may explain the depleted values in their hair, and obscure any interspecific difference in foraging height.

Alternatively, carbon values may indeed reflect an overlap in the forest strata used for foraging activities. Although C. satanas on Bioko Island were typically observed in higher strata, feeding activities occurred primarily in the middle and lower canopy, as did those of C. erythrotis (Gonzalez-Kirchner 1996, 1997). Dietary analysis of M. leucophaeus in lowland forests of Bioko showed that the fruit of tree species to make up the overwhelming majority of their diets in this habitat (Chapter 2). In addition, the diets of C. satanas are primarily composed of seeds, not leaves (Gautier-Hion et al. 1997;

Tutin et al. 1997), which have carbon isotope values similar to fruits (O’Leary 1981;

Carter 2001; Cerling et al. 2004; Sandberg et al. 2012). Therefore the similarity in carbon isotope values between the primate species might be a result of a lack of vertical partitioning of foraging activities. Foraging observations made during this study are

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consistent with this explanation; however empirical dietary data on C. satanas or C. erythrotis from Bioko Island are needed to confirm this hypothesis.

Differences in the nitrogen isotopes resulted in two distinct groups, with C. erythrotis and P. monticola significantly higher than C. satanas and M. leucophaeus.

Insects are known to be an important part of the diets of guenons, second only to fruits

(Glenn & Cords 2002), which is consistent with the enriched δ15N values of C. erythrotis.

The high δ15N of P. monticola on Bioko Island is similar to duikers (including P.

monticola) in LuiKotale, Democratic Republic of Congo. There three species of duikers

had remarkably similar δ15N values to the primate species (including bonobos,

mangabeys, and colobus) within the same forest (Oelze et al. 2011). The authors contend that, similar to δ13C values, this reflects the similarly high fruit consumption of duikers and primates. However, the significant difference between C. erythrotis and M. leucophaeus are contrary to this explanation, given that the primary dietary component of both these species is fruit. Some legumes have been shown to deplete the δ15N of

consumers, however as none were identified in the diet of M. leucophaeus from fecal

sample analysis or observations (Chapter 2), consumption is likely too low to

significantly vary their δ15N values. Insect remains comprised 2.4% of the dry fecal

sample weight of M. leucophaeus on Bioko Island (Chapter 2), and observations of insect consumption are rare in C. satanas in Lope Reserve, Gabon, making up only 2.6% of all feeding observations (Tutin et al. 1997).

Given this information, we argue that the nitrogen isotope values in all species, including P. monticola, are indeed reflective of trophic level. Animal matter, including insects and carrion, is regularly reported in the diets of P. monticola, and active hunting

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of ants and Diptera have been observed in this, and other, duiker species (Dubost 1984;

Wilson 2001). It is also possible that the fruits consumed by duikers contain increased

numbers of insects, as they are predominantly those which have ripened to the point of

falling from trees, or are discarded by primates during foraging activities. Piles of

dropped fruit under ripe Ficus vogeli in lowland forests on Bioko Island are often

teeming with adult and larval insects (pers. Obs.), which may result in considerable

inadvertent consumption by foraging duikers. Regardless of the mechanism of insect

consumption, no other sources of variation in nitrogen isotopes explain these differences.

Sexual Differences

The lack of significant difference in the δ13C and δ15N values between the sex

classes of any species in our study is similar to the findings of previous studies of wild

primates (Schoeninger et al. 1997, 1998; Smith et al. 2010; Oelze et al. 2011). However

the significantly reduced nearest neighbor distance values, and relatively smooth density

plots of females of all species indicates that they have a more specialized diet than males,

with less individual variation. Intraspecific dietary differences between sex classes may

be a result of differing nutritional requirements and morphological constraints related to

body size and reproductive role. The adult body mass of male monkeys is often greater

than females, as is true for all monkeys on Bioko Island (Butynski et al. 2009), resulting

in higher overall metabolic requirements for males. Pregnancy, lactation, parental care,

and gamete production in females increase demand for energy, proteins, and mineral

(Kerr 1972; Gautier-Hion 1980; Jolly 1985a; Rose 1994), and may require unique

foraging strategies to meet these demands.

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Among sexually dimorphic species, morphological differences may allow specializations in foraging, dietary composition, and differentiate niches between sex

classes (Jolly 1985a). For example, compared to females, male capuchins (Cebus

capuchinus) in Santa Rosa National Park, Costa Rica, spent more time on the ground,

used larger and lower branches to forage on, were in proximity to members of the same

sex less, and were better able to access prey in difficult to access locations (under bark)

because of their increased body weight, strength, canine teeth and jaws (Rose 1994).

Females concentrated their foraging efforts on foods with low nutritional return on

foraging effort, but that provided the most consistently reliable return, such as insect

larvae or bird eggs, whereas males used a more opportunistic foraging strategy, taking a

wider assortment of commonly available foods with a higher risk of complete loss of

investment, such as large invertebrates and vertebrates (Rose 1994). Such sexual

differences in foraging strategies would likely result in isotope niche patterns found in

this study, and may ultimately reduce competitive interactions between sexes.

Seasonal Differences

Seasonal variations in rainfall and moisture that occur on Bioko Island are

expected to result in predictable seasonal fluctuations in the isotopes values in plant

tissues throughout the year. However, the lack of seasonal differences in the mean values of the any consumers does not support this prediction. Instead it implies that there is little

seasonal variation in basal resources, and that consumers do not shift to alternative

resources or altitudinal ranges throughout the year, as has been reported in bonobos (Pan

paniscus) (Oelze et al. 2011), but contrary to mountain gorillas (Gorilla beringei)

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(Blumenthal et al. 2012). An alternative explanation may be that shifts in consumption

coincide with changes in the isotope values of plants. However, as the hair samples were

not subsampled, the most likely explanation for the seasonal similarity is that the hair

samples contained dietary data for both wet and dry seasons.

However, reduction of the niche area and individual dispersal in the niche space

(as indicated by NDD) in M. leucophaeus suggest they focus on a more narrow isotopic niche in the wet season. Given that on average, 90% of their fecal sample weight in the dry season was composed of fruit, it is likely that the niche reduction denotes an even higher consumption of fruit during the wet season, as was reported of M. leucophaeus in

Korup National Park, Cameroon (Astaras 2009). Further analysis is needed to determine

if seasonal differences do exist.

Niche Overlap and Competition

There was a considerable amount of intraspecific variation in the isotopic niches

of all species. This was particularly true of P. monticola and C. satanas; contrary to

dietary information derived from populations on mainland Africa that indicated they

would have the more specialized diets of the four species (e.g(Dubost 1984; Glenn &

Cords 2002; Astaras 2009). Both P. monticola and C. satanas populations had individuals with δ15N values above 8 ‰, which are indicative of high insect consumption,

while others ranged between a spectrum of diets with varying levels of omnivory,

frugivory, and folivory (Schoeninger et al. 1999). In stark contrast to the highly

omnivorous individuals, three C. satanas were below 3 ‰, which reflect an absence of

insect consumption, and heavy consumption of fruit or leaves of leguminous species.

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Due to this large separation between conspecific individuals in the δ-space, there

was substantial overlap in the total niche space covered by each species. These results indicate that the potential for interspecific resource competition may be high. However, the lack of identifiable species-wide strategies to avoid this competition suggests that

resources may not be limited beyond their demand. Given the substantial individual variation found in isotope values, if interspecific competition were imposing high selective pressure on one or more species, we should expect to find populations shifting towards alternative strategies (Bolnick et al. 2003). In contrast, the large isotope niches of

P. monticola and C. satanas are in line with the expectations of niche expansion into open niche space resulting from competitive release (Diamond 1970a, 1970b). These findings are contrary to predictions based on the species-area curve, which suggest that primate species richness on Bioko Island is considerably greater than its area is able to support (Cowlishaw 1999).

Carcass Origins

Stable isotopes have been used to track animal movements and identify the geographic origins of animal tissues in a variety of taxa (Rubenstein & Hobson 2004).

However these studies have predominantly used heavy elements (e.g. Strontium, and

Sulfur) that are extremely costly and, in many cases, inappropriate to the study scope or geographic region of interest, or tissues that are invasive or difficult to collect (e.g. bone collagen or enamel apatite). Recently, an increasing number of ecologists have highlighted the benefits of developing tracking techniques from stable isotopes of light elements (e.g. δ13C, δ15N, δ18O, δ2H) (e.g. Rubenstein & Hobson, 2004; West et al., 2006;

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Schoeninger, 2010). These benefits include the low costs of services from a large number

of facilities that are able to quickly perform the analyses, and the ability to analyze tissues

such as feces and hair, which require minimally invasive collection methods. Such

methods typically rely on dietary information to infer sample origins, and have been

successfully employed to track a wide diversity of taxa (Hobson 1999).

One of the underlying objectives of this study was to determine if the ecological

information gained during this study would provide some insight into the forest origins of

the hair samples analyzed. Unfortunately, the high intraspecific variation we found is

likely to inhibit fine-scale locality information. However, the δ13C values of M.

leucophaeus do help eliminate some areas.

Organisms that consume coastal crabs or other marine-derived foods would be

13 expected to have tissues enriched in δ C, similar to values of those consuming C4 grasses

(Crowley 2012). Fecal sample analysis of M. leucophaeus on Bioko (Chapter 2) found that the consumption of African land crabs (Johngarthia weileri) was unique to

individuals living in the island’s southeastern coast. On the southwestern coast, where

there was no apparent difference in crab availability, there was no evidence of crab

consumption from field observation or fecal samples (N = 81). In addition, C4 grasses

were a regular food item within the Gran Caldera, at elevations above 900 m asl., but were never recorded in the feces or foraging observations at any other location on the island. Because of these dietary differences, if hair samples originated from individuals living in either the southeastern coast or the Gran Caldera we would expect to find much higher δ13C values. This suggests it is unlikely that any of the drills sampled were hunted

in either of these locations. However, there is no way to confirm this prediction without

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reference samples from that location, or increased information on the spatial variations in

basal resources across the island.

Conclusions

Based on these findings, we conclude that while the potential for resource

competition between C. erythrotis, P. monticola, C. satanas, and M. leucophaeus on

Bioko Island might be high, there is currently little evidence of niche shifts to alleviate

competitive interactions. Instead, differences between the predicted and realized isotope

niches of P. monticola and C. satanas suggest they may have expanded to fill open niche

spaces resulting from the extirpations of 16 species following the island’s separation from

the mainland. The resulting individual variability in isotope values of these two species

reflects a wide spectrum of dietary strategies and trophic positions. In contrast, M.

leucophaeus and C. erythrotis were found to have relatively small isotopic niches that are

indicative of primarily frugivorous diets, and increased omnivory in the latter species.

Females of each species in the study were shown to have reduced isotopic niches

compared to males; however no consistent trend was found between the wet and dry

seasons. Differences between the sex classes were only apparent in respect to their

combined δ13C - δ15N values; none existed when the isotopes were analyzed

independently. As demonstrated in these results, the inclusion of such community or population wide niche metrics can reveal important ecological differences that would otherwise be missed by univariate comparisons. To date, most isotope ecology studies have yet to apply these techniques, despite their utility.

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High individual variability in the isotope values of both the sources and consumers limited the ability of the mixing model (SIAR) to develop strong predictions about the dietary contributions of food sources. In general, C3 plants are the primary food

resource of all four species, while no species consumed C4 grasses. This strongly

suggests that hunting activities are primarily occurring outside of the Gran Caldera and

the southeastern coastal forests. These results indicate that it is unlikely that isotope

analysis will be able to identify the forest origins of primate bushmeat carcasses at a fine

spatial scale on Bioko Island using only nitrogen and carbon isotopes. Future work would

benefit from increased sampling in a systematic pattern that would provide isotope values

for a greater diversity of source species, across the entire southern third of Bioko Island.

The results of this study have important implications to the potential use of hair

samples derived from bushmeat carcasses. Intra– and interspecific niche comparisons can

provide information that is vital to our understanding of broader ecological, behavioral,

or evolutionary questions. However, this information is often difficult and time

consuming to collect using traditional methodologies, such as observations and fecal

sample analysis. Stable isotopes provide a tool in which ecological information can be

rapidly collected and analyzed on individual species, or entire ecological communities.

To our knowledge, this is the first stable isotope study using samples entirely derived

from bushmeat carcasses. The findings of this study have provided valuable information

on the ecology Bioko Island primates, of which little information was previously

available. Based on these results, we argue that the collection of samples can be

performed in a manner which does not promote unsustainable bushmeat hunting, but

provides information important to conservation programs.

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13 15 Figure 5.1: Boxplots of δ C and δ N isotopes (top) and total %C and %N (bottom) for C3 plants, C4 grasses, and invertebrate food sources.

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Figure 5.2: Relationship between the carbon isotope values 13 (δ C‰) of herbaceous plant tissues and elevation.

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13 15 Figure 5.3: Boxplots of the δ C‰ (left) and δ N‰ (right) values of consumer hair samples.

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Figure 5.4: Minimum convex polygons of each species and their sex classes in the 13 15 δ C and δ N niche space. Total area occupied (“δ space”) by the polygons and nearest neighbor values (mean +SEM) are provided.

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13 15 Figure 5.5: Plots of the δ C and δ N values for each species. Points correspond to each individual sampled with their sex class denoted by color; those with error bars (SD) represent the means. Kernel density estimates of the points are shown as contour lines (middle plots) for the niche space biplots, and separately for the corresponding 13 15 frequency distributions of the δ C and δ N (bordering plots).

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Figure 5.6: Stable isotope values of consumer hair (symbols: individual means corrected by the TEF values) and source tissues (symbols: mean + SD) plotted in the 13 15 δ C - δ N niche space.

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Figure 5.7: SIAR boxplots of the predicted proportions of each food source to the diets of the consumer species (boxes represent the 95, 75, and 25% credibility intervals).

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Table 5.1: Summary results for the analysis of food sources, including the total nitrogen and carbon of the tissues and their stable nitrogen and carbon values. Total % N Total % C Food Type N δ15N ‰ (SD) δ13C ‰ (SD) (SD) (SD) Aframomum spp. 13 0.7 (0.36) 0.54 (2.07) 42.92 (1.73) -30.85 (2.69) C3 34 1.26 (0.79) 1.11 (2.14) 43.29 (3.73) -30.51 (2.64) C4 3 1.73 (0.89) 1.24 (0.97) 41.9 (0.48) -11.36 (0.46) Fruit 7 1.44 (0.53) 1.18 (2.2) 46.49 (5.86) -28.71 (1.42) Ground Cover 4 2.38 (0.49) 3.56 (1.47) 39.52 (1.74) -34.18 (0.71) Invertebrate 6 10.76 (0.97) 5.32 (1.0) 47.19 (1.82) -27.93 (1.26) Leaf 8 1.63 (0.81) 1.35 (1.46) 43.58 (2.17) -31.55 (1.82) Root 7 1.41 (0.88) 1.23 (1.79) 41.91 (2.42) -31.02 (2.73) Stem 11 0.83 (0.66) 0.91 (2.7) 42.28 (2.0) -31.05 (2.69) Tree 16 1.47 (0.73) 1.02 (1.97) 44.78 (4.43) -29.65 (1.68)

Table 5.2: Results of the ANOVA's and subsequent multiple comparisons (Tukey's Post Hoc Test, p value) of the δ13C‰ (top) and δ15N‰ (bottom) values of food sources. δ13C ‰

ANOVA: F5,80=117.37, p< 0.001 Aframomum spp. C4 Grass Fruit Ground Cover Invertebrate Leaf Root Stem Aframomum spp.       C4 Grass <0.001        Fruit  <0.001       Ground Cover <0.001 <0.001       Invertebrate <0.001 <0.001  <0.001     Leaf  <0.001   <0.001    Root  <0.001   0.008    Stem  <0.001   0.002   Tree  <0.001  <0.001     δ15N‰

ANOVA: F5,80=14.12, p< 0.001 Aframomum spp. C4 Grass Fruit Ground Cover Invertebrate Leaf Root Stem Aframomum spp.       C4 Grass         Fruit         Ground Cover 0.002        Invertebrate <0.001 <0.001 <0.001      Leaf     <0.001    Root     <0.001    Stem     <0.001   Tree    0.01 <0.001   

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Table 5.3: Summary information for the stable isotope analysis of consumer hair samples. δ13C ‰ δ15N ‰ Species Sex Season N mean SD mean SD Dry 5 -24.65 0.92 5.34 0.3 Female Wet 5 -24.44 0.6 4.99 0.46 M. leucophaeus Dry 5 -24.72 0.5 5.13 0.73 Male Wet 6 -25.52 1.1 4.72 0.87 Dry 5 -24.47 0.76 6.52 0.68 Female Wet 6 -24.58 0.73 6.0 1.13 C. erythrotis Dry 5 -24.79 1.42 5.82 0.37 Male Wet 5 -24.65 0.75 6.17 1.58 Dry 10 -25.35 1.09 4.47 0.95 Female Wet 1 -23.8 NA 5.84 NA C. satanas Dry 5 -25.26 0.82 5.18 1.71 Male Wet 5 -24.61 1.24 4.09 1.36 Female Dry 5 -25.57 0.54 6.81 1.12 P. monticola Male Dry 5 -26.03 0.99 6.28 1.6

Table 5.4: Total isotope niche space (δ13C-δ15N) as determined by the area of the minimum convex hulls occupied by each species, their sex, and the season in which the hair samples were collected. All Total Samples Wet Dry Species Samples Wet Dry Male Female Male Female Male Female M. leucophaeus 6.3 5.4 2.06 5.0 2.0 3.47 0.72 0.81 0.81 C. erythrotis 9.18 5.56 4.05 7.36 4.16 3.43 3.22 1.53 1.67 C. satanas 15.95 6.15 9.27 6.4 3.27 4.1 na 2.25 2.79 P. monticola 4.68 na 4.68 3.33 1.54 na na 3.33 1.54 Total 36.18 17.56 19.17 15.47 11.4 11.77 4.09 11.02 10.51

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155

CHAPTER 6: DISSERTATION CONCLUSIONS AND BROADER

IMPLICATIONS

Synopsis of the Dissertation

The drill has been an enigmatic species since its initial description by western

scientists over 200 years ago. Very few ecological or behavioral studies have been

performed in the wild because of their elusive nature and dense rainforest habitats,

leaving much of their natural history poorly understood. This is particularly true of the

Bioko Island drill, which had never been empirically studied, prior to the study presented

herein. The purpose of this dissertation was to fill the major gaps that exist in our

understanding of the fundamental aspects of the life of Bioko Island drills. In addition, by

comparing this with the better studied mainland drills and mandrills we gain a more

comprehensive understanding of the genus Mandrillus, and the extent to which local

environmental conditions vary their ecology and behavior. The Gran Caldera and

Southern Highlands Reserve in southern Bioko Island provided an ideal site to perform

this investigation, as significant differences in the habitat structure and composition exist

between montane and lowland forests within a relatively small area, and with a

contiguous population of drills. Here I will summarize the major findings of each of the

studies presented in chapters 2-5, review their limitations and future research directions, and discuss the conservation implications of this work.

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Feeding Ecology

In chapter two I used primary and secondary feeding observations, fecal sample

analysis, and food availability assessments to compare the qualitative and quantitative

dietary differences that exist between drills in lowland and montane forests. Drills

exhibited two significantly different dietary patterns related to the contrasting

availabilities of fruits and herbaceous vegetation between these habitats. In the lowland

forest, where fruits were in relatively high abundance, drills were primarily frugivorous.

They consumed the fruits of numerous tree species, but showed an affinity for certain

species. This diet is consistent with what has been reported of mainland drills and

mandrills (Hoshino 1985; Astaras 2009).

In contrast, likely due to fruit scarcity in the montane forest, drills consumed a

primarily folivorous diet composed of the stem-pith and roots of terrestrial herbaceous

plants. Plants of the genus Aframomum were the primary food item of these drills. This

diet was also marked by a significant increase in the consumption of insects, mushrooms,

and leaves. As members of the Cercocebus -Mandrillus clade of Papionin monkeys, drills

are thought to be uniquely adapted to exploiting seeds during periods of fruit scarcity

(Fleagle & McGraw 1999, 2002). Studies of mainland drills and mandrills have found the

consumption of seeds to increase significantly during seasonal fruit shortages, providing

support to this argument (Hoshino 1985; Astaras 2009). However, Bioko Island drills do

not exhibit this strategy in relation to spatial variations in fruit availability, and the

fallback diet they consume has never been previously reported in studies of Mandrillus

spp.

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Drills were omnivorous at all three study sites, consuming at least 18 species of

invertebrates and two vertebrates. However, between sites there were conspicuous

differences in the species eaten. Some of these differences might be explained by

variations in availability, such as the regular predation of African giant millipedes

(Archispirostreptus gigas) in the Caldera, and others that may not, particularly in the

consumption of African land crabs (Johngarthia weileri) at Moaba Playa, but not at

Moraka Playa. We found no evidence of predation on larger vertebrates, such as duikers,

despite previous reports suggesting that this may occur (Schaaf et al. 1990; Gonzalez-

Kirchner & de la Maza 1996).

Gastrointestinal Parasitic Infections

In chapter three I analyzed fecal samples to determine in differences in the species

richness or prevalence of infections existed between drills in montane and lowland

forests. We also sought to determine if individual dietary differences were related to

individual variations in parasite species presence or richness. There were no significant

differences or relationships found in any of the analyses performed. Drills in both forest

types were infected by species common to other primates, with the exception of

Cyclospora papionis. This is the first study to report this coccidian species outside of

olive baboons (Papio anubis) in east Africa, representing a considerable expansion of its

range (Eberhard et al. n.d., 2001). Parasite richness and prevalences were within a range expected from other primate parasite analyses.

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Group Size, Polyspecific Associations, and Habitat Use

In chapter 4 I used data collected during group encounters made from transects

and blinds to estimate the average size and composition of Bioko Island drill groups, as

well as the rate they are found in polyspecific associations with other monkeys. I also

performed a habitat assessment along transects in the montane and lowland forests to

determine if differences existed in their habitat use between habitats. No significant

differences existed in the group sizes or structure between forest types, and there were

only minor differences found in the estimates from each survey type. The proportion of

encounters with polyspecific associations was low, and observations suggest that they are

likely random, infrequent occurrences. Analyses of the habitat associations were largely

unsuccessful at finding any significant relationships between the locations of group

encounters and the habitat at that point. However, there was an indication that drill

encounters in montane forests were more evenly dispersed within their habitat, and that

drills in the lowland forest use certain locations at higher frequencies than others. This is

consistent with the dispersal patterns of their primary food items in each habitat.

Groups were remarkably smaller than any previous quantitative analyses of

Mandrillus groups. The average group sizes of drills reported from studies using similar

methods were between 52.3 and 93.1 individuals per group, and groups were composed

of a minimum of five to a maximum of 400 individuals (Gartlan 1970; Wild et al. 2005;

Astaras et al. 2008). In contrast, the mean group size of Bioko Island drills was less than

five individuals per group, and only one group of 20 individuals was encountered during

the study. Due to this extremely low group size, the average composition of groups on

Bioko Island was also considerably different than previous reports of the Mandrillus. The

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ratio of adult males to all other individuals was 1:4 in groups of Bioko drills, and we

found evidence of the occurrence of numerous single-male/single-female groups.

Mainland drills and mandrills are typically reported to have male ratios of 1:20 for all

group members, and between 1:5 and 1:12 for females. Previous studies have found that in response to high interspecific competition, an absence of natural predators, and intense

human hunting activities, primate groups may benefit from having fewer individuals,

eventually leading to persistent monogamy (Tilson 1977; Watanabe 1981). This is remarkably similar to the current situation on Bioko Island, indicating that the differences

between Bioko drills and mainland Mandrillus may, indeed, reflect ecological variations.

Stable Isotope Ecology

In chapter 5 I used the stable isotope analysis to assess the potential competition

between drills, red-eared guenons, black colobus, and blue duikers, and identify any

strategies they may use to mitigate its effects. The isotopic niches of all four species

overlapped considerably, indicating the potential for resource competition may be high.

Females in all species had smaller and more densely distributed isotope niches, which is

consistent with the findings of many observational/behavioral primate studies (Gautier-

Hion 1980; Jolly 1985b; Rose 1994). Substantial intraspecific variation in the isotope

niches existed among the blue duikers and black colobus, resulting in these two species

having the largest isotope niches of the four studied. Blue duikers and black colobus also

had individuals with isotope values indicative of high levels of omnivory; both of these

results were contrary to expectations. The findings of this study suggest that drills and

red-eared monkeys may primarily consume fruits, with little variation among individuals,

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but that the colobus and duiker may be expanding into open niche space existing on the

island. The diet of the drill indicated by the stable isotope analysis is consistent with the

diet of drills in the lowland forest.

Limitations and Future Directions

The findings of this research contrast some of the longstanding assumptions of the

ecology and behavior of the drill, namely in their group size and composition, and

feeding behaviors. These differences warrant further investigation, and would benefit

particularly from several lines of investigation. There are also aspects of the design or

implementation of the research presented in this dissertation that might be improved to

better clarify the results.

1.The dietary analysis we performed only included fecal samples collected during the dry

season because of the difficulty of finding samples and the time expenditure it would

require. However, even a few samples collected during the wet season would help

determine if the trends found in this study hold true between seasons, and if drills in

lowland forests consume seeds as a seasonal fallback food.

2.Differences in the availability, distribution, and nutritional quality of the food items

consumed are likely to explain the dietary differences we found between forest types. A

more comprehensive habitat assessment that included survey plots throughout both the

montane and lowland forests, and provided high resolution data on the species

abundance, spatial distribution, phenology, and biomass of vegetation would enable this

to be tested. Analysis of the nutritional content of Aframomum, pre- and post-

consumption would facilitate the modelling of foraging strategies.

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3.The parasite analysis was performed on only 51 samples, and from only two sites. An

increase in sample size may result in the detection of additional parasite species, and

also improve the power of the statistical analyses. As these samples were collected

from unknown individuals, molecular analysis of the fecal samples to determine the

identities and sexes of the individuals would address any overlap in the sample set, and

also provide additional hypotheses to test. The identification of Cyclospora papionis in

drills raises several important questions: 1) how and when did it get to Bioko Island, 2)

what other species does it infect, 3) are other Cyclospora spp. present in the

community? These questions have important implications to the spread and

diversification of humans, and should be addressed.

4.The group size and composition of drills we reported here were primarily from foraging

groups. Few were counted from sleeping sites, which are more likely to contain the

entire social group, if Bioko drills do divide into daily foraging subgroup. The

observations made during this study provide some small indications that daily

subdivisions may occur in some areas, namely Moaba Playa, but not in others. Because

of the high hunting pressure, no attempts to habituate drill groups should be made until

adequate enforcement of the hunting ban is realized. GPS and radio-telemetry would

not only help address this question, but could be used to improve on every aspect of this

study. Because telemetry would enable groups to be quickly located, fecal samples

could be easily collected during the wet season during the intermittent periods without

rain, and the sample sizes of each of these studies could be improved in considerably

less time. Finally, telemetry would provide information concerning the altitudinal

movements of drills.

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Conservation Implications

Drills have been identified as one of the most threatened primates in Africa due to

their highly restricted distribution and taxonomic distinction, and the intensity of the

threats of shotgun hunting and habitat loss which they face, (Gadsby & Jenkins 1997;

Grubb et al. 2003). Ultimately, their continued survival is largely dependent on two

factors: 1) their adaptability to environmental variation, and 2) the efficacy of

conservation actions. It is clear that drills have high ecological and behavioral plasticity.

The dietary plasticity found in Bioko Island drills is among the most dramatic of any

primates in the world. However, the rate at which they are being killed for the

commercial bushmeat trade is clearly beyond the sustainable rate. In 2006, Hearn,

Morra, & Butynski estimated that only 3000-4000 drills remained on Bioko Island. In the last year alone, an estimated 1753 drills were sold at Semu bushmeat market (D. Cronin, personal communication, November, 2013). Although bushmeat is known to be imported from the mainland of Cameroon, these carcasses are smoked (dried) before being transported by boat to Bioko Island (M. K. Gonder, personal communication, December

4, 2013), smoked carcasses only account for 12.6% of those sold at the market (Cronin

2013). Based on these estimates, it is likely that the remaining population of Bioko Island drills is only a fraction of the 2006 estimate, and could be well under 1,000 individuals.

The development of new roads in the Gran Caldera and Southern Highlands

Scientific Reserve is certain to exacerbate the already poor conservation outlook for

drills. The GCSH likely represents the only effective population on the island, and are

thus critically important to the survival of the subspecies. There is no doubt that, unless

the existing bans on primate hunting are enforced, the increased accessibility to the

163

southern beaches and the crater of the Gran Caldera will quickly decimate the remaining

populations of drills and the other highly endangered species, especially the red colobus.

The importance of protecting the GCSH cannot be overstated, and I recommend that

immediate steps be taken to limit access, and enforce hunting bans. Several other authors

have made recommendations on the steps necessary to conserve drills and other primates

throughout their range (Butynski & Koster 1994; Castroviejo et al. 1994; Gadsby &

Jenkins 1997; Wild et al. 2005; Hearn et al. 2006; Astaras 2009; Cronin et al. 2010;

Cronin 2013; Morgan et al. 2013), which I will reiterate.

1. Bioko Island’s protected areas need delineation and protection. There are few

roads accessing these areas, and checkpoints already exist at multiple points on

route from the market to the forest. If the soldiers manning these checkpoints

were properly trained and mandated to enforce the hunting bans, hunting would

quickly be reduced.

2. The use of dogs in hunting drills is devastating to their populations, as large

proportions or entire groups can be removed from the forests during each hunt.

Dogs should be banned and removed from both protected areas immediately.

3. Shotguns should be confiscated from all residents.

164

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VITA

Jacob R. Owens 3245 Chestnut Street, PISB 503 Philadelphia, PA 19104 [email protected]

EDUCATION Ph.D., Environmental Science, Drexel University, Philadelphia, PA 2013 B.S., Biology, The Richard Stockton College of New Jersey, Galloway, NJ 2006

PROFESSIONAL EXPERIENCE Research Associate, Bioko Biodiversity Protection Program (BBPP) 2008-2013 Logistics and Volunteer Coordinator, BBPP Gran Caldera Expedition 2008-2013 Field Course Instructor, Drexel University 2012-2013 Teaching Assistant, Drexel University 2010-2013 Research Assistant, The Wetlands Institute 2006-2007

AWARDS AND GRANTS Drexel University Graduate Studies’ International Travel Award 2012 Drexel University Biology Department Travel Award 2012 Primate Conservation Incorporated Research Grants 2009, 2010, 2011

SELECTED PUBLICATIONS AND ABSTRACTS Eberhard, M. L., J. R. Owens, H. S. Bishop, M. E. de Almeida, A. J. da Silva, G. W. Hearn, and S. Honarvar. Cyclospora from Drills (Mandrillus leucophaeus poensis) on Bioko Island, Equatorial Guinea (in press). Emerging infectious diseases. Cronin, D.T., J. R. Owens, H. Choi, S. Hromada, R. Malhotra, F. Roser, R. A. Bergl, Where has all our research gone? A 20-year assessment of the peer-reviewed wildlife conservation literature (in press). International journal of comparative psychology. Owens, J. R., S. Honarvar, and G. W. Hearn, Integrating dietary and intestinal parasite data to improve the conservation strategies of the Bioko Island drill (Mandrillus leucophaeus poensis), International Congress for Conservation Owens, J. R., S. Honarvar, and G. W. Hearn, Altitudinal Variation in the Feeding Strategies of the Bioko Island Drill (Mandrillus leucophaeus poensis), XXIV Congress of the International Primatological Society Owens, J. R., and Hearn, G. W., Conservation Status of the Bioko Island Drill (Mandrillus leucophaeus poensis), XXIII Congress of the International Primatological Society