DISSERTATION

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RESEARCH PROJECT – BIOL3005
POTENTIAL ANTHROPOGENIC EFFECTS ON TH HEALTH AND
PHYSIOLOGICAL STRESS OF THE HONDURAN PALEATE SPINY-TAILED IGUANA
(CTENOSAURA MELANOSTERNA)
Guillaume Demare
guillaume.demare.09@ucl.ac.uk
University College London
UNIVERSITY SUPERVISOR: PROF ROGER WOTTON
FIELD SUPERVISOR: DR CHAD M. MONTGOMERY

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TABLE OF CONTENT
1. LIST OF ABBREVIATIONS...................................................................................5
2. ABSTRACT............................................................................................................6
3. INTRODUCTION....................................................................................................7
3.1.Anthropogenic impact on biodiversity ........................................................7
3.2.The Honduran paleate spiny-tailed iguanas................................................7
3.3.Indirect impact of humans on animal populations .....................................8
3.4.Implications of physiological stress for conservation.............................10
3.5.Measuring health .........................................................................................11
3.6.Project aims and hypotheses .....................................................................12
4. MATERIALS AND METHODS.............................................................................13
4.1.Description of study site .............................................................................13
4.1.1. Geographical features ..........................................................................13
4.1.2. Collection sites .....................................................................................15
4.2.Capture methods .........................................................................................17
4.2.1. Capture.................................................................................................17
4.2.2. Marking.................................................................................................18
4.3.Collection of blood samples for analysis ..................................................19
4.3.1. Blood sampling methods ......................................................................19
4.3.2. Processing of blood samples................................................................19
4.4.Measurements..............................................................................................20
4.5.Statistical methods ......................................................................................21

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5. RESULTS.............................................................................................................22
Preliminary results.......................................................................................22
5.1.Health indicators and variation due to sex and life stage........................25
5.1.1. Variation in health indicators because of sex effect .............................25
5.1.2. Variation in health indicators due to life stage ......................................25
5.2.Interrelationship between health indicators..............................................28
5.3.Health and anthropogenic disturbance .....................................................30
5.3.1. Variation in health indicators between sites .........................................30
5.3.2. Health indicators and capture status ....................................................32
5.4.Glucocorticoid steroid hormones: expected results................................33
6. DISCUSSION .......................................................................................................34
6.1.Health indicators and variation ..................................................................34
6.1.1. Variation due to sex and life stage .......................................................34
6.1.2. Health and external parasitism .............................................................36
6.2.Health indicators and anthropogenic disturbance ...................................37
6.2.1. Anthropogenic disturbance does not affect health condition ................37
6.2.2. Potential evidence of acclimation to human disturbance......................38
6.2.3. Prospects for stress physiology and human disturbance .....................39
6.3.Implications and future directions .............................................................41
7. ACKNOWLEDGMENTS ......................................................................................42
8. REFERENCES.....................................................................................................43
9. ANNEXES ............................................................................................................48

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1. LIST OF ABBREVIATIONS
ANCOVA: Analysis of covariance
ANOVA: Analysis of variance
BCI: Body condition index
GC: Glucocorticoid steroid hormone
HCRF: Honduran Coral Reef Foundation
HCT: Haematocrit
HPA: Hypothalamic–pituitary–adrenal axis
IUCN: International Union for Conservation of Nature
N: sample or population size
NPF: North Palm Forest (collection site)
P/A: Presence/Absence
PIT-tag: Passive Integrated Transponder tagging
RS: Research Station (collection site)
SD: Standard deviation
SE: Standard error of the mean
SSD: Sexual size dimorphism
(Sub)Adult: grouping of subadults and adults individuals
SVL: Snout-to-vent length

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2. ABSTRACT
A rising concern that accompanies the increasing need to protect biodiversity on the
long-term is the poorly investigated indirect effect of human activities, such as nature-
based tourism, that potentially jeopardise the persistence of animal populations
already threatened by direct anthropogenic pressure. The present study explores the
potentially negative effects of human disturbance on the health and stress physiology
of an endangered species of iguana. To date, there has been relatively little
investigation linking anthropogenic disturbance and the health of free-living animals.
As part of the study, the health of spiny-tailed iguanas (Ctenosaura melanosterna)
was assessed, based on body condition, haematocrit level and external parasite
load. The effect of anthropogenic disturbance was determined by comparing the
health of individuals frequently interacting with humans with that of individuals
thriving in a natural environment. The results indicate that human disturbance does
not affect health condition of iguanas, which is possibly mediated through a
behavioural adjustment allowing them to invest less time and energy in predator
avoidance. Acclimation has been observed on several instances but the long-term
effect on fitness (in terms of survival and reproductive output) has never been tested.
If data on stress physiology confirms that spiny-tailed iguanas have the ability to
acclimate to human disturbance, the population studied would then offer the rare
opportunity to explore the effect of acclimation on long-term fitness. This would in
turn have important implications for the management of animal populations
threatened by human activity.

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3. INTRODUCTION
3.1.Anthropogenic impact on biodiversity
The deleterious anthropogenic impact on biodiversity is now well recognised.
Humans have caused a dramatic increase in the rate of species extinctions, and this
realisation has come with the need to identify areas and species worth protecting
(Pimm et al., 1995). The gradual loss of the natural capital has often been attributed
to the direct impact that humans have on the environment through expansion of their
activity. The consequent habitat loss is often considered to be the prime factor
accounting for current rates of biodiversity loss (Kerr and Currie, 1995).
While this trend is true for most taxa, long-term studies on reptiles have brought
evidence for a global decline, resulting from both anthropogenic and natural causes
(e.g. diseases). Humans have a significant impact on reptiles, mainly through habitat
degradation and destruction, introduction of invasive species, environmental
pollution, unsustainable use, and anthropogenic climate change (Gibbons et al.,
2000). Whilst the latter is the object of much debating, climate change has
nonetheless been shown to be a significant threat to reptile communities (e.g.
Pounds, Fogden and Campbell, 1999).
3.2.The Honduran paleate spiny-tailed iguana, Ctenosaura melanosterna
The Honduran paleate spiny-tailed iguana (Ctenosaura melanosterna) is a good
example of a reptile species whose populations have declined because of direct
anthropogenic pressure. It is a member of the Iguanidae family and was formerly
considered to be synonymous with Ctenosaura palearis, until its recent description as
a separate species (Buckley and Axtell, 1997). The species is endemic to Honduras
and is classified as Endangered by the IUCN due to its limited distribution,
fragmented populations, habitat destruction, and illegal harvesting for food and
international pet trade. The species is currently estimated at fewer than 5000 mature
individuals (Pasachnik, Montgomery and Henningheim, 2011). The Ctenosaura
genus includes other closely related species that are also endangered because of
direct anthropogenic impact, including C. palearis, C. bakeri and C. oedirhina
(Gutsche and Köhler, 2008; Stewart et al., 2011). The latter two species are both

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insular (located on the islands Utila and Roatan, respectively) and endemic to
Honduras (Pasachnik, 2006).
Spiny-tailed iguanas are diurnal, omnivorous and semi-arboreal. Individuals generally
feed on leaves, fruits and flowers, and will opportunistically feed on small vertebrates
and invertebrates, including carrion (Blázquez and Rodríguez-Estrella, 2007).
The distribution of C. melanosterna is limited to one population in the subtropical dry
forests of the Valle del Aguán on the mainland, and two populations in the Cayos
Cochinos Archipelago off of the north coast of Honduras. The Cayos Cochinos,
located within a biological reserve, contains the highest density of individuals with the
majority inhabiting Cayo Menor (Pasachnik, Montgomery and Henningheim, 2011).
The biological reserve provides protection to the marine and terrestrial wildlife as part
of a management plan implemented by the Honduran Coral Reef Foundation
(HCRF). It insures continuous monitoring and maintenance of biodiversity, its
sustainable use by local communities (Garifuna populations), and provides support
for scientific research and community development (HCRF, 2008). Cayo Menor is
exclusively used for the HCRF research station, thus inhabited only by employees
(including researchers) and visitors (e.g. research volunteers). Therefore, the
population most sheltered from direct anthropogenic impact is the one located on
Cayo Menor.
3.3.Indirect impact of humans on animal populations
Indirect anthropogenic impact, as used here, refers to any human factor that has a
direct impact at the individual level, but which overall effect on the population is only
detectable on the long-term. However, the dichotomy between direct and indirect
impact is often difficult to establish.
One field in conservation biology that takes indirect anthropogenic disturbance into
consideration when managing animal populations is reintroduction biology.
Translocation of animals from a source to a recipient population is a common
practice in wildlife management that is employed in order to reinforce a declining
unsustainable population. This approach to conservation is delicate and is only
effective if individuals successfully establish and persist in the novel habitat. While
habitat quality and population dynamics are regarded as major factors determining

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translocation success, other factors such as the intrinsic condition of the animal being
relocated are increasingly viewed as crucial components. This includes general body
condition, health state, and physiological stress conditions (Armstrong and Seddon,
2007). Translocation generally involves capture, handling, captivity, transport and
release to novel site, which altogether represent a sequence of stressful events,
inducing consecutive and long-lasting acute stress responses in the animal. In most
cases, this leads to chronic stress in the relocated animal. Chronic stress can
subsequently translate into starvation, increased susceptibility to disease, reduced
reproductive capacity, and reduced ability to adequately respond to predation. These
effects collectively reduce the individual’s fitness, and ultimately, the overall
probability of translocation success (Dickens, Delehanty and Romero, 2010).
This area of conservation biology puts forward the potential threat represented by
anthropogenic disturbance on animal populations. Understanding the indirect
detrimental effects that humans have on an animal’s overall fitness is important in
order to successfully manage populations that are already threatened by direct
human impact. Consequently, the attempt to fully protect endangered animal
populations puts a question mark on human activities such as ecotourism, which fly
the flag of biodiversity whilst potentially bringing heavy disturbance to protected
animals. However, relatively few studies have explored the negative indirect effects
of ecotourism on reptiles. Frequent interaction with humans potentially stress and
drive animals to invest considerable amounts of time and energy into behavioural
avoidance (Buckley, 2004). Similarly, ongoing research on endangered populations,
which usually involves captures, handling, confinement and processing of animals,
can potentially bring similar levels of detrimental disturbance.
These activities, primarily aiming to protect wildlife on the long-term, should therefore
go past simple assumptions and incorporate the indirect effect of anthropogenic
disturbance into practice. The present study aims to assess the effect of such
disturbance on the health of an endangered species.
Indirect anthropogenic impact includes direct disturbance, as well as modification of
the environment. For example, unintentionally providing animals with an
inappropriate diet can result in reduced health. The importance of adequate
nutritional resources has indeed been suggested for the conservation of various
reptile species (Oftedal and Allen, 1996). While the present study focuses on the

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general effect of humans on health condition, it also intends to investigate the effect
of anthropogenic disturbance on stress physiology.
3.4.Implications of physiological stress for conservation
Appropriate response to environmental stimuli is essential for survival. In vertebrates,
physiological and behavioural coping with environmental stressors is mainly realised
through the hypothalamic–pituitary–adrenal (HPA) axis, in the form of release of
glucocorticoid steroid hormones (GC). The primary role of these hormones is to
regulate the overall energy in the body, including mobilising energy for specific
responses to an environmental ‘stressor’ (Busch and Hayward, 2009). It is generally
accepted that a good stress response is characterised by a low baseline GC level, a
fast acute GC response, and rapid triggering of subsequent negative feedback. The
acute release of GC is vital, as it induces multiple responses that help animals
survive life-threatening events, including mobilisation of glucose resources and the
‘fight-or-flight’ response (Breuner, Patterson and Hahn, 2008). Moreover, the release
of GC seems to mediate the trade-off between the costs associated with reproduction
and survival: environmental factors inducing a GC release generally promote survival
over reproduction, although this is known to be context-dependent (Moore and
Jessup, 2002; Breuner, Patterson and Hahn, 2008).
However, the continuous stimulation of the HPA axis results in ‘chronic stress’, which
in turn leads to health defects and reduced fitness. Chronically stressed individuals,
as mentioned previously, become unable to adequately cope with environmental
stressors, which in turn dramatically reduces their chances of survival. Alternatively,
animals can avoid the immediate detrimental effects of chronic stress by acclimating
to the repeated stressor (e.g. human disturbance). On the other hand, this is known
to modify the response to novel stressor, which can potentially jeopardise fitness on
the long-term (Busch and Hayward, 2009, Romero, 2004).
Physiological stress response is therefore closely associated with general health
condition, and can be potentially useful as a biomarker to reflect anthropogenic
disturbance (Romero, 2004).

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3.5.Measuring health condition
The fitness of an individual largely depends on body condition and energy availability.
Using the residuals from the linear regression of body mass with respect to a body
size indicator is believed to be the most widely used method for estimating body
condition (termed body condition index, BCI) of vertebrates (Green, 2001). Although
this method has been criticised on several instances, it has been shown to generally
follow the basic assumptions of linearity, independence between energy reserves
(e.g. fat reserves) and body size, and independence between true body condition
and body size (Schulte-Hostedde et al., 2005).
This method is indeed extensively used for determining the body condition of reptiles,
using snout-to-vent length (SVL) as a body size indicator (e.g. Dunlap and Mathies,
1993).
The level of haematocrit (HCT), sometimes measured as packed cell volume, is also
often used as a health indicator (e.g. Komoroske et al., 2011). It directly measures
the proportion of erythrocytes in the blood, and therefore reflects the potential of the
animal’s oxygen-carrying capacity. There is a strong positive relationship between
haemoglobin content and size of erythrocytes. Reptiles have fewer but larger
erythrocytes compared to birds and mammals, but also have an overall lower amount
of haemoglobin per unit of blood (Hawkey et al., 1991). This is likely to reflect the
low-oxygen demanding lifestyle associated with ectothermy. Reptiles maintain a
lower level of HCT (about 29% in lizards), but it is important that this level is kept
constant in order to maximise oxygen-carrying capacity and blood flow rate, which
together optimise delivery of oxygen throughout the body. Thus, deviation from
optimum HCT level (in either direction) is likely to have subsequent negative
consequences on fitness (Pough, 1980).
External parasite load (e.g. ticks, mites) generally reflects immune function and can
therefore be used as a health indicator. Indeed, reptiles that mobilise more energy in
other activities have been observed to suffer from a reduced immune function, which
in turn translates into higher ectoparasite load (Schall, Prendeville and Hanley,
2000). Furthermore, external parasitism relates to the health indicators described
above. Increased number of ectoparasites can reduce both haematocrit level and

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body condition (Dunlap and Mathies, 1993). In fact, studies where ticks are
experimentally removed have shown that higher tick load was associated with lower
overall fitness (Main and Bull, 2000).
3.6.Project aims and hypotheses
The primary aim of this research study is to assess the effect of anthropogenic
disturbance on the health (based on the indicators body condition index, haematocrit
level, and external parasite load) and physiological stress (measured as the
corticosteroid hormone baseline level and acute response) of Ctenosaura
melanosterna. Secondary aims include evaluating the level of variation caused by
sex and life stage in the different health indicators used, in corticosteroid hormone
levels (baseline and response), as well as investigating the relationship between the
different health indicators used, and between health and physiological stress.
From this, the following hypotheses can be identified:
H1 There is no difference in health condition between anthropogenically-disturbed
iguanas and iguanas found in a natural environment.
H2 There is no difference in GC levels between anthropogenically-disturbed
iguanas and iguanas found in a natural environment.
H3 Sex and life stage do not cause any variation in health condition, or in GC
levels.
H4 There is no correlation between the different health indicators used.
H5 There is no correlation between health and physiological stress.

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4. MATERIALS AND METHODS
4.1.Description of study site
4.1.1. Geographical features
The study was conducted on Cayo Menor, the second largest island (covering 0.64
km2
of land area) in the Cayos Cochinos archipelago (15°58’18.99”N,
86°28’31.34”W) located off of the north coast of Honduras, 39 km northeast of La
Ceiba. The archipelago also comprises a larger island, Cayo Grande (1.55 km2
), as
well as several (13) small cays, and is situated within the boundaries of a biological
reserve known as the ‘Reserva Biológica Cayos Cochinos’ (Figure 1). Cayo Menor
consists of a central North-South ridge reaching a maximum elevation of 141 m. Its
vegetation is mainly composed of oak forest (about half of the island canopy is
dominated by tropical lowland oak Quercus oleoides), mature mixed forest on some
sheltered slopes (including the canopy tree Pouteria sp. and understory tree Roupala
montana), and mixed secondary forest (e.g. the deciduous tree Bursera simarouba)
(Bermingham et al. 1998).
In addition to forest, the island also consists of four major herpetofaunal habitats:
coconut palm groves, mangrove forests, rocky promontories and hill forests. The
latter two are known to provide habitat for Ctenosaura melanosterna (Wilson and
Cruz Diaz 1993). The herpetofauna of Cayo Menor consists of 16 documented
species, including 1 anuran (Similisca baudinii), 9 lizards (e.g. Anolis allisoni,
Ctenosaura melanosterna, Cnemidophorus lemniscatus) and 6 snakes (e.g. Boa
constrictor, Dryadophis melanolomus) (Wilson and Cruz Diaz 1993; McCranie Wilson
and Köhler, 2005).

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4.1.2. Collection Sites
I collected individuals at two different sites on the island: (1) within and around the
research station (RS), and (2) at the north palm forest (NPF) (Figure 2).
The RS compound is located at the south end of the island and includes sleeping
facilities, a dry laboratory, and a restaurant. It is the only portion of the field site that
exhibits anthropogenic disturbance to C. melanosterna, including indirect contact with
the numerous volunteers and staff, direct contact due to ongoing research, artificial
habitat provided by the station infrastructures and maintenance of a helicopter
landing pad, and artificial diet due to food waste on which iguanas have been
observed to feed.
The NPF consists of the narrow region at the north tip of the island, and is mainly
characterised by relatively low palm groves alternating with sandy substrate. The
NPF was chosen as a collection site as it contains numerous individuals, and is
situated relatively far away from the RS. Thus, individuals were never found to travel
from one collection site to the other.
The difference in environmental conditions between the two sites offers the
opportunity to test the effect of anthropogenic disturbance on the health of C.
melanosterna, which is the central aim of the present study.

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Figure 2 Map showing the locations of the two chosen study sites – North Palm
Forest (NPF) and Research Station (RS) – on Cayo Menor.
Photos: Daniel Nicholson (NPF) and HCRF (available at
http://www.cayoscochinos.org) (RS).

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4.2.Capture methods
4.2.1. Captures
Iguanas were located during random walks across the two sites, and caught on an
opportunistic basis (capture methods are described in Table 1).
In total, 126 individuals were collected at the RS, and 25 individuals at the NPF. This
gap in sample size accounts for the difficulties associated with capturing iguanas in
wooded areas rather than in open areas (common at RS), as well as capture
opportunities.
Table 1 Description of the capture techniques employed.
Description Use
Stationary Net
10m long/1m high net extended
transversally on the ground.
Iguanas moving freely on the
ground eventually entangle in the
net and become immobilised.
Nets are checked every 30 min
for captures.
To catch individuals
moving at ground level.
The size of the net
mesh selects for adult-
sized iguanas.
Noose
2-3 m long pole (wood or metal)
to which a piece of wire is fixed at
one end into a running knot. The
noose is typically about 10-20
cm, 5-10 cm, and 3-5 cm
diameter for trapping adults,
juveniles, and hatchlings,
respectively. Captures entail
placing the noose around the
neck region of the iguana and
gently pulling the pole upwards
To catch individuals on
the ground or resting in
trees.
Adjusting the diameter
of the noose allows
selection for any age-
class.
Hand-catch
Typically involving about 3
volunteers that surround an
individual in order to catch it by
hand. Precautions include
grabbing the individual behind the
head to avoid bites, as well as
securing the legs to avoid
scratches.
Generally for catching
individuals on the
ground, in open spaces
(i.e. difficult to apply in
wooded areas).
This technique can be
used for catching
individuals of any size.

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4.2.2. Marking
Prior to release, each individual was marked using the following methods. First, a
PIT-tag (Passive Integrated Transponder) was inserted subcutaneously, left of the
vertebrae on the posterior-dorsal side, using a hypodermic syringe. This provides
each individual with a permanent and unique identification code that can be read
upon recapture using a PIT-tag reader (type Biomark). Secondly, a bead-tag was
placed below the rear nuchal crest spines and provides individuals with a unique
code of 1-3 coloured beads (see Binns and Burton, 2007). While the beads allow
external identification (i.e. without catching/handling), they have been observed to be
lost because of fighting or mating events (i.e. not permanent). The combination of
those two methods is used for marking adults, subadults and juveniles (Figure 3A,
3B).
Because of their small size (about 6.0 cm SVL), neither PIT- nor bead- tagging could
be used for hatchlings. Instead, they were marked using toe-clips, which can provide
a number for each hatchling (front left: 1-5; front right: 6-10; hind left: 20-60; hind
right: 70-110; maximum two toes, excluding the longest toe of the hind limbs – i.e.
toes 50 and 100).
Finally, all individuals were applied with white marking on both flanks in order to
distinguish recently caught iguanas and avoid repeated captures (Figure 3C).
However, this method is only temporary as the paint wears off after the next ecdysis
event.
Figure 3 Internal and external marking techniques used. A: PIT-tags and hypodermic
syringe; B: bead-tag (e.g. ‘dark green/dark blue’); C: white marking.

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4.3.Collection of blood samples for analysis
4.3.1. Blood sampling methods
Blood samples (about 0.3 ml each) were drawn either from the postorbital sinus
using heparinised capillary tubes (length 75 ±0.5 mm, I.D. 1.1-1.2 mm), or from the
caudal vein using heparinised syringes (1.0 ±0.01 ml). The first blood draw was
performed immediately upon capture in order to obtain an estimate of the GC
baseline level. The second blood draw was done after the animal was placed in a
bag for 30 min, which simulates a continuous stressful event. The final draw was
therefore used to estimate the GC stress response of each individual.
4.3.2. Processing of blood samples
After the blood samples were stored at low temperature for less than 12 hours, they
were processed for different purposes. First, a sample extract was placed in a
capillary tube and spun at high speed (8000 rpm) for 5 min using a Readacrit
centrifuge. The haematocrit (HCT) level (the portion of the blood accounting for red
blood cells in %) was recorded using a microhaematocrit capillary tube reader (±1%).
The remaining portion of the sample was then centrifuged at 6000 rpm for 5 min, in
order to extract the plasma that contains the corticosteroid hormones.
Once the plasma was extracted from a sample, it was placed in a vial and stored at
- 20°C and shipped to Virginia Tech to be analyzed by Dr Ignacio Moore.

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4.4.Measurements
Following the final blood draw, individuals were inspected for the external parasites
ticks and mites, both found attached to the skin surface. Despite varying dramatically
in size (from mm to cm), an accurate tick count was possible. However, mites are
found in clusters of tiny individuals, allowing only an account for presence/absence.
Several measurements were also made: snout-to-vent length (SVL) (using a
standard measuring tape; ±1 mm), body mass (using an electronic scale; ±0.1 g),
and cloacal temperature (taken upon capture using a thermometer type T
Thermocouple; ±0.1°C). The body condition index (BCI) of each individual was
determined as the residual of the relationship of logSVL against logMass.
Then, the sex of each individual was determined through direct observation of the
femoral pores that males exhibit along the ventral side of the hind limbs (absent in
females) (Figure 4), and confirmed through cloacal probing by inserting a steel probe
of adequate diameter inside the vent and towards the tail. In males, the probe
reaches further posteriorly as it passes between the hemipenes. Hatchlings were
considered too small for reliable sex determination and were therefore not assigned a
sex-class.
The age-class (hatchling, juvenile, and subadult/adult) was deduced based on SVL
(cm): SVL > 18.5 cm for (sub)adults; SVL = 9.2-15.3 cm for juveniles; SVL < 7 cm for
hatchlings.
Figure 4 The femoral pores (e.g. red arrows) are present in male iguanas on the
ventral side of the hind limbs (A), but are absent in females (B).

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4.5.Statistical methods
The variation in health indicators (BCI, HCT, ectoparasite load) caused by sex and
life stage was explored using a combination of t-tests and one-way ANOVA tests.
When the outcome of ANOVA was a statistically significant difference, a Tuckey-
Kramer test was used in order to account for the difference. The relationship between
ectoparasite load and body size was initially investigated using a linear regression
analysis. The variation in ectoparasites due to sex and life stage was further
analysed using a general linear model ANCOVA with body size (SVL) as a covariate.
The interrelationship between health indicators was explored through separate paired
comparisons, using t-tests and linear regression analyses.
Finally, a series of t-tests and chi-square tests were performed to test for site and
capture status (recapture, new capture) differences in body condition, haematocrit
level, tick number, and mite presence/absence.
All statistical analyses were carried out using the software JMP version 9 (SAS
Institute Inc., Cary, NC, USA, 1989-2007), and a significance level of 95%.
The data on GC levels was not available at the time of the statistical analysis, and
was therefore excluded.

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5. RESULTS
The principal aim of the research is to assess the differences in health condition
between anthropogenically-disturbed iguanas (at the RS) and iguanas found in a
natural environment (at the NPF). The health was assessed according to three
different indicators: body condition index (BCI), haematocrit (HCT) level, and external
parasite load (tick number and ratio of mite presence/absence).
Preliminary results
The relationship between mass (g) and SVL (cm) was used in order to obtain an
estimate of the body condition index (BCI) for each individual. This relationship is
exponential and is required to be log-transformed in order to obtain linearity (Figure
5). The BCI is calculated as the residual of logMass, so that any data point above
and below the line of best fit corresponds to a positive and negative BCI,
respectively.
Then, a t-test was used to analyse the difference in mean SVL (cm) between
(sub)adult males and females. The results indicate that males are on average larger
than females (Figure 6A), with significant statistical difference (t48 = 4.49; P <
0.0001). Thus, based on the data used in this study, it is possible to conclude that C.
melanosterna shows sexual size dimorphism (SSD) at adulthood.
The graph in Figure 6B shows the difference in SVL (cm) between the different age
classes. Iguanas that have a SVL above 18.5 cm were categorised as (sub)adults,
those in the range of 9.2-15.3 cm SVL were described as juveniles, and any iguana
measuring below 7 cm SVL were considered hatchlings.

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Figure 5 Relationship between body mass (g) and SVL (cm). A: Each data point
corresponds to the body mass with respect to SVL of each iguana collected and
measured (N = 147). B: The data was log-transformed to obtain linearity. The slope
of the line of best fit for the relationship ‘logMass by logSVL’ is of +3.14.
The green, red and blue data points correspond to hatchlings, females and males,
respectively (unknown sex is labelled black).
Figure 6 SVL (cm) with respect to: (A) sex in (sub)adults; (B) life stage (hatchlings,
juveniles and (sub)adults). The data points correspond to the SVL of each measured
individual. A: (sub)adult males and females have a mean SVL ± SE of 28.5 ± 0.7 cm
and 24.7 ± 0.5 cm, respectively. B: The range in SVL (mean ± SE) is: 18.5-35.5 cm
for (sub)adults (26.1 ± 0.3 cm); 9.2-15.3 cm for juveniles (11.3 ± 0.5); 5.2-7 cm for
hatchlings (6.0 ± 0.2).

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Finally, the data collected on cloacal temperature (i.e. body temperature, °C) reveals
that individuals maintain a range of 31.2-38.6°C, with a mean temperature (± SE) of
34.4 ± 0.2°C (Figure 7). Iguanas are ectotherms and regulate their body temperature
depending on environmental conditions (e.g. weather, sun exposure, time of day).
Without controlling environmental factors, temperature is therefore not expected to
correlate with health indicators, which do not fluctuate in time at the same pace. On
the other hand, temperature data can be used for comparison with corticosteroid
hormone acute stress response, as the speed of the response may vary depending
on the body temperature that the animal exhibits during the stressful event.
Figure 7 Distribution of cloacal temperature (°C) among sampled individuals (N =
34). Each bar represents the number of individuals located within a 0.5°C
temperature interval, with standard error bars. The box plot above the graph
indicates the value of the mean ± SE (34.4 ± 0.2°C), the median (34.25°C), the lower
(33.6°C) and upper (35.6°C) quartiles, and the minimum (31.2°C) and maximum
(38.6°C) values.

25
5.1.Health indicators and variation due to sex and life stage
5.1.1. Variation in health indicators because of sex effect
The first research aim is to explore the variation in health condition due to sex effect,
based on 3 different indicators. A series of t-tests were carried out to compare BCI,
HCT level, and tick load between males and females. The fitness of ratio in mite P/A
was tested using a chi-square test. The tests were performed by combining
individuals from both the RS and NPF. Detailed analyses are shown in Annexe 1.
The results indicate that there is no statistical difference in mean BCI between males
and females (t59 = 0.22; P = 0.8287). There is also no difference in mean HCT level
(t32 = -1.27; P = 0.2132). However, there is a significant difference in mean tick
number between males and females (t59 = 2.23; P = 0.0298), with males presenting
more ticks on average. Finally the results of the chi-square tests indicate that there is
no difference in ratio of mite P/A between males and females (χ2
= 0.044; P =
0.8339).
From this we can conclude that sex does not seem to affect health condition, based
on the health indicators BCI, HCT and mites, but does affect the number of ticks
present on the skin of an iguana. Males have on average (± SD) 8.6 ± 2.4 ticks,
whereas females only have 2.1 ± 1.6 ticks.
5.1.2. Variation in health indicators due to life stage
The second research aim is to test the effect of life stage on health condition, using
the same indicators. A series of ANOVA tests were employed in order to identify any
difference in BCI, HCT level and tick number between hatchlings, juveniles and
(sub)adults. In the case of significant difference at the 0.05 confidence level, a
Tuckey-Kramer test helped identifying where the difference lied. The fitness of ratio
in mite P/A was assessed using a chi-square test. Detailed analyses are presented in
Annexe 1.
The results show that there is no significant statistical difference in mean BCI
between the different age classes (F144, 2 = 0.34; P = 0.715). There is also no
difference in mean HCT (F37, 2 = 0.62; P = 0.5432). However, there is a significant
difference in tick load (F2, 144 = 7.88; P = 0.0006), with (sub)adults exhibiting more

26
ticks than hatchlings (P = 0.0005). The chi-square tests showed that there is also a
difference in the ratio of mite P/A (χ2
= 6.231; P = 0.0444), with juveniles and
(sub)adults having mites more often than expected, and hatchlings less often than
expected.
Therefore, juveniles and (sub)adults have on average more ticks and present mites
more often than hatchlings.
The results from the last two sections suggest that external parasite load differs
between males and females (tick number), and between hatchlings, juveniles and
(sub)adults (tick number and mite presence). Thus, it seems that larger (or older)
iguanas present a higher level of external parasitism, compared to those that are
smaller (or younger).
These observations lead to test whether SVL (i.e. body size) is an important factor for
determining external parasite load. First, the relationship between SVL and tick load
(Figure 8.A) was tested using a linear regression analysis. Secondly, a t-test was
used to explore the difference in SVL between iguanas with and without mites
(Figure 8.B). The two tests indicate that there is a significant positive correlation
between SVL and tick number (r2
= 0.115; t145 = 4.35; P < 0.0001), and that larger
individuals present mites significantly more often than smaller individuals do (t145 =
2.36; P = 0.0194).
In addition, because the positive correlation between tick load and SVL seems to be
driven by the few outliers (see Figure 8), the regression analysis between SVL and
tick number was also done excluding those points. The result was still a significant
positive correlation (r2
= 0.202; t141 = 5.97; P < 0.0001).

27
Figure 8 Relationship between SVL (cm) and external parasite load – tick number
(A); mite P/A (B). A: The slope of the line of best fit for the relationship ‘tick number
by SVL’ is of +0.26. The graph also displays the 95% confidence intervals for the fit.
B: Individuals that are not infected by mites have a mean SVL ± SE of 12.4 ± 0.8 cm,
whereas infected individuals have a mean SVL ± SE of 17.3 ± 1.9 cm.
The green, red and blue data points correspond to hatchlings, females and males,
respectively (unknown sex is labelled black).
Finally, ANCOVA tests were used in order to account for the variation caused by
body size (SVL), to test whether the difference in external parasite load is partly due
to sex and life stage. The results indicate that life stage has no statistically significant
effect on tick load (F = 0.89; P = 0.4132) or ratio of mite P/A (χ2
= 1.57; P = 0.4565).
Sex also has no effect on tick number (F = 2.28; P = 0.1367).
From this we can conclude that neither sex nor life stage have a significant effect on
any of the health indicators used, including external parasite load, which seems to
depend largely on body size (SVL). As a consequence males, females, hatchlings,
juveniles and (sub)adults can be grouped together for subsequent analyses involving
any health indicator.

28
5.2.Interrelationships between health indicators
As part of the study, it is important to test the effect of one health indicator over
another. The aim is to determine the relationships between health indicators through
paired comparisons using: (1) regression analyses to compare two numeric variables
(HCT, BCI and tick number); (2) t-tests to compare a numeric variable with a nominal
variable (mite P/A).
A series of 3 regression analyses and 3 t-tests allowed every combination of
indicators to be tested (illustrated in Figure 9).
The results (detailed in Annexe 2) indicate that there is no significant correlation
between HCT level and BCI (r2
= 0.039; t37 = 1.22; P = 0.2289), between tick load
and HCT level (r2
= 0.0005; t38 = -0.14; P = 0.8893), or between tick number and BCI
(r2
= 0.003; t143 = 0.71; P = 0.4814). There is also no difference in HCT level between
individuals with mites and those without mites (t38 = -0.82; P = 0.418), in BCI (t143 =
0.13; P = 0.8991), or in tick number (t145 = 0.05; P = 0.9603).
Thus, it is possible to conclude that the health indicators used are independent from
one another. This means that if an individual is found to be unhealthy based on one
indicator, the same does not necessarily apply based on a different indicator.

29
Figure 9 Relationship between paired health indicators, using data from every
sampled iguana on Cayo Menor. A: HCT level by BCI (slope = +10.9); B: tick number
by HCT level (slope = -0.05); C: tick number by BCI (slope = +6.2); D: HCT level by
mite P/A; E: BCI by mite P/A; F: tick number by mite P/A. The graphs A, B, C display
the line of best fit as well as the 95% confidence lines. The graphs D, E, F, show the
mean and standard deviation for each group of individuals (horizontal green lines).

30
5.3.Health and anthropogenic disturbance
Iguanas that are assumed to be anthropogenically disturbed were compared with
those that are suspected to be less affected by such disturbance.
First, the health of iguanas was compared between the two sites, based on the
indicators previously used. Secondly, newly captured iguanas were compared with
iguanas that had been captured in the past (i.e. recaptured during this study).
Recaptured iguanas are supposedly more anthropogenically disturbed, as the
capture event is considered to be stressful, and can potentially affect health on the
long term.
5.3.1. Variation in health indicators between sites
First, iguanas from the RS were compared with iguanas from the NPF (illustrated in
Figure 10). A series of t-tests and a chi-square test allowed identifying the difference
in mean health level between the two groups of individuals.
The results, detailed in Annexe 3, reveal that iguanas from the RS do not deviate in
health condition from iguanas found at the NPF. There is statistically no difference for
BCI (t145 = -0.11; P = 0.9096), HCT level (t38 = 0.33; P = 0.7424), tick count (t145 =
1.32; P = 0.1876), and ratio of mite P/A (χ2
= 0.002; P = 0.9612).
Thus, iguanas living in a human disturbed environment do not seem to deviate in
health indicators from iguanas living in natural conditions.

31
Figure 10 Health level of iguanas from the RS and from the NPF, for each health
indicator. A: BCI by site; B: HCT level by site; C: tick number by site; D: count of
individuals with (dark blue) and without (light blue) mites within each site (with
percentage displayed above each bar, for individual site). The graphs A, B and C
also display the mean and standard deviation (horizontal green lines).

32
5.3.2. Health indicators and capture status
The health indicators were then compared according to status of capture. Iguanas
that were captured at least once in the past (during a previous study) and recaptured
in this study were compared with newly captured iguanas (illustrated in Figure 11),
using a series of t-test and a chi-square test. It is important to note that newly
hatched individuals were excluded from the analysis, as they represented a bias in
‘new captures’.
The results, detailed in Annexe 3, indicate that there is statistically no difference in
HCT (t35 = 0.21; P = 0.8372), tick number (t62 = 0.56; P = 0.5776), and ratio of mite
P/A (χ2
= 0.558; P = 0.455) between recaptured and newly captured iguanas.
However, there is statistical difference in BCI (t62 = 2.68; P = 0.0095). Newly captured
iguanas have a mean BCI ± SE of -0.03 ± 0.01, whereas recaptured iguanas have a
mean BCI ± SE of 0.02 ± 0.01.
Therefore, recaptured iguanas present a higher body mass with respect to their body
size (measured as SVL), compared to newly captured iguanas.

33
Figure 11 Health level of newly captured and recaptured iguanas, for each health
indicator. A: BCI; B: HCT level (%); C: tick number; D: count of newly captured and
recaptured individuals with (dark blue) and without (light blue) mites (with percentage
displayed above each bar, for each status of capture). The graphs A, B and C also
display the mean and standard deviation (horizontal green lines).
5.4.Glucocorticoid steroid hormones: expected results
The stress hormone data is still being processed by Dr Ignacio Moore, and is
therefore not available as of yet. Nevertheless, stress physiology will be discussed in
the context of the results obtained above.

34
6. DISCUSSION
6.1.Health indicators and variation
6.1.1. Variation due to sex and life stage
The results detailed in the previous section indicate that neither sex nor life stage
have an effect on body condition (measured as BCI) or haematocrit level. Thus,
based on these indicators we can infer that individuals have similar energy and
oxygen requirements for daily activity, regardless of age or sex (Peterson, 2002).
However, males are found to have significantly more ticks than females. This
difference could result from a sexual variation in behaviour. If one sex is
characterised by larger territory areas and greater moving distances, then external
parasite load can increase because of higher opportunity for parasite attachment
(Godfrey et al., 2010; Aubret et al., 2005). Other studies have identified that a
differential cost in reproduction between males and females (e.g. production of
testosterone in males) results in a difference in immune function (i.e. trade-off
between costs of reproduction and immune system). This in turn leads to contrasting
levels of external parasitism between sexes (Moller, Sorci and Erritzoe, 1998;
Salvador et al., 1996). A difference in external parasite infection (both ticks and
mites) is also seen between individuals of separate life stage: (sub)adults and
juveniles both show a higher load of ectoparasites compared to hatchlings. Similarly
to sex, behavioural differences between hatchlings and older individuals could result
in variations in opportunity for parasite attachment and host immune function,
thereby accounting for the results obtained.
The results also suggest a significant positive correlation between body size (SVL)
and ectoparasite load. After removing the effect of body size, neither sex nor life
stage have an effect on external parasite load. This shows that the difference
observed is not due to intrinsic disparities between males and females, or between
iguanas of different life stage, but is largely due to the body size of the individual.
Sexual size dimorphism (SSD) (with males larger than females) and the dramatic
increase in body size from hatchling to adulthood therefore translate into differential

35
patterns of ectoparasitism occurrence in iguanas. Effectively, a greater skin surface
area would increase the likelihood for parasite attachment (i.e. higher opportunity).
Moreover, ticks and mites are known to select specific areas on the host for
attachment and subsequent feeding (Nelson et al., 1975). Although no statistical
analysis was carried out to compare attachment sites, the general trend is that ticks
were localised in sheltered areas on the iguana: at the base of the limbs, in the
nuchal area, and within skin folds. Ticks have a selective advantage in choosing
these sites because it minimizes risks of dislodgment associated with the host’s
mechanical disturbance (e.g. when moving) (Nelson et al., 1975). In addition, these
areas on the host have been shown to correspond to sites of higher vascularisation
in spiny-tailed iguanas, providing a feeding advantage for the parasites (Weathers
and Morgareidge, 1971). As a consequence, crowding effect might be an important
factor in limiting number of external parasites on smaller-sized individuals. However,
inference about parasite competition is difficult without clear species identification of
the parasites observed.
While body size seems to account for most of the variation observed in external
parasitism, it is possible that sexual differences in external parasitism are only
detectable seasonally. Iguanas were inspected well after the breeding season (in
July-August), which occurs during the spring. Since ticks and mites usually attach for
less than a month, as observed in other iguanid species (with variations depending
on parasite-host association and environmental factors such as precipitation and
climatic cycles), the differential in ectoparasite load resulting from sexual differences
may have been undetectable at the time of inspection (Goldberg and Bursey, 1991).
It should also be noted that no difference in mite infection is observed between
males and females, which is likely to be due to the lack of resolution in counting
mites (only presence/absence) combined with the fact that the difference in body
size is relatively small between males and females.
Finally, because parasite load seems to be mostly the consequence of variations in
body size, it may be a poor indicator for immune function in the population studied.
Nevertheless, it is crucial to identify the relationship between external parasite load
and health condition, and the potential negative impact it has on the C. melanosterna
population.

36
6.1.2. Health and external parasitism
The negative effect of external parasitism has been subject to much investigation in
free-living populations. Higher load of ectoparasites lowers both body condition and
haematocrit levels of wild birds, which can have negative consequences on
metabolic capacity and survivorship (Merino, Mínguez and Belliure, 1999; Simon et
al., 2004). Similar results were found in reptile populations, as mentioned previously.
Infestation with ticks lowers haematocrit level, and when coupled with malarial
infection (e.g. Plasmodium mexicanum), body condition is also reduced (Dunlap and
Mathies, 1993). Most importantly, studies experimentally removing external parasites
have showed that tick infestation could reduce overall fitness, as it was clear that
individuals with higher tick load had smaller home range sizes, reduced speed and
lower endurance (Main and Bull, 2000).
However, the results obtained in the present study show that levels of external
parasitism do not correlate with any other health indicators. Thus, infection by ticks
and mites seems to have no significant effect on body condition or aerobic capacity
(haematocrit level), indicating that external parasitism has little or no effect on the
fitness of the studied population. In fact, it has been suggested that parasitism could
have no obvious negative effects at low levels of infection, but may become
detrimental above a certain threshold (Dunlap and Mathies, 1993). On the other
hand, even the few individuals that were found with an abnormally large number of
ticks (up to 60), had HCT levels and BCI within healthy ranges. The effect of higher
tick load may have been undetected at the time of inspection because the ticks on
those individuals were generally small, indicating recent attachment, and located in
places where future dislodgment was likely (i.e. high load may be a transient phase).
Finally, other environmental factors can mask the negative effect of elevated
parasitism. For example, high habitat quality can help maintain a good body
condition, thereby creating a balance with the costs associated with elevated
parasitism (Main and Bull, 2000). Therefore, it is possible that iguanas with higher
external parasitism modulate their behaviour in order to compensate for the costs of
having more ticks and mites. This issue could be further tested by controlling
environmental variables such as habitat quality and feeding patterns of iguanas with
varying levels of parasitism.

37
6.2.Health indicators and anthropogenic disturbance
6.2.1. Anthropogenic disturbance does not affect health condition
Predator avoidance is a costly behaviour because it mobilises both time and energy
that could otherwise be invested towards other activities such as foraging, and has
been shown to cause long-term negative effects on fitness (e.g. reduced growth,
Downes, 2001; reduced body condition, Martín and López, 1999). However, the
effect of increased behavioural avoidance resulting from anthropogenic disturbance
has rarely been investigated. A study carried out on the wall lizard (Podarcis muralis)
showed that nature-based tourism, as a form of human disturbance, has a negative
effect on body condition and immune function (Amo, López and Martín, 2006). While
lizards located in areas with heavy tourism showed similar antipredatory behaviour
compared to individuals less exposed to disturbance, they performed predatory
avoidance more often. As a result, those individuals had lower body condition,
reduced immune function, and higher levels of parasitism. Hence, it was suggested
that human disturbance is energy demanding and can therefore have adverse
consequences on overall fitness (Amo, López and Martín, 2006).
However, the present study did not find any evidence of negative consequences on
health resulting from anthropogenic disturbance. No significant differences in body
condition, haematocrit level or ectoparasite load were identified between iguanas
living in a human-disturbed site and those living in a natural environment. This
indicates that the current research, which can be perceived as a form of tourism,
does not seem to have negative effects on the health of spiny-tailed iguanas, based
on the indicators used in this study. From this we can infer that iguanas are not
affected by human disturbance, either because they have acclimated to humans, or
because the RS actually presents significantly lower levels of disturbance compared
to urban areas and tourism. In both cases, iguanas would invest relatively little
amounts of time and energy in behavioural avoidance, thereby maintaining good
body condition and competent immune function.
However, the present study relies on the major assumption that all iguanas from the
RS are equally affected by human disturbance. Thus, the potential effect of
disturbance may be masked in this analysis if there were significant variations in
human disturbance among individuals collected at the RS compound.

38
Complementing the results with data on physiological stress will allow us to identify
whether the assumption was complied with, as GC levels can be used as a
biomarker for disturbance (Romero, 2004).
An additional factor that might affect the comparison between the two sites is the
relatively low sample size collected at the NPF (N = 25).
6.2.2. Potential evidence of acclimation to human disturbance
The final aspect of the study was to compare iguanas that had already been
captured in the past with those that were newly captured. While the two groups did
not differ in haematocrit level and external parasitism, suggesting similar aerobic
capacities and immune competency, they had different body conditions. Iguanas that
were recaptured were found in better body condition, indicating that they have better
energy reserves and higher relative fitness (Schulte-Hostedde et al., 2005).
This finding is consistent with a semi-natural experiment carried out on Iberian wall
lizards (Podarcis hispanica), which investigated the effect of non-lethal predation
(e.g. human disturbance) on body condition (Rodríguez-Prieto, Martín and
Fernández-Juricic, 2010). Individuals that regularly encounter a low-risk predator
were found to adjust their antipredatory behaviours (e.g. fleeing) in order to
maximise energy investments for survival. As a result, these individuals could not
only allocate more time and energy in foraging, they could also reduce the costs
associated with accumulative avoidance behaviour. Acclimation to non-lethal
predators can therefore be adaptive because it allows individuals to increase relative
fitness, as suggested by the increase in body condition (Rodríguez-Prieto, Martín
and Fernández-Juricic, 2010). The process of acclimation therefore offers a
seductive explanation for the difference in body condition observed between
recaptured and newly captured iguanas. Individuals that undergo a first capture may
‘learn’ that humans can be regarded as non-lethal predators. Avoidance behaviour is
subsequently adjusted in order to allocate more energy in other activities, thereby
increasing body condition and fitness. Without the experience of first capture, other
iguanas fail to acclimate to human disturbance to the same extent.
On the other hand, the process of acclimation has been suggested to promote
habituation to the wrong predator (Frid and Dill, 2002). Thus, acclimation may be
adaptive but potentially lead to the new cost of inadequately lowering vigilance and

39
avoidance behaviour. This issue is especially relevant in the case of C.
melanosterna as it is still largely threatened by poaching (Pasachnik, Montgomery
and Henningheim, 2011). If individuals are found to acclimate to humans, they could
become unable to adequately respond to poachers, as well as to natural predators
(e.g. Boa constrictor on Cayo Menor).
Several methods could allow us to test the hypothesis that recaptured iguanas follow
a process of acclimation. First, the newly captured iguanas could be later recaptured
and analysed to see whether body condition has increased, as expected if
acclimation occurs. Secondly, we could investigate the patterns in GC levels.
Human-induced stress response, as discussed below, should indicate whether
iguanas are acclimated to human disturbance, or whether an alternative hypothesis
should be considered.
An alternative explanation for the observed difference between recaptured and newly
captured individuals is a sampling bias towards individuals with higher body
condition. Past captures may have been concentrated in areas where individuals
have easier access to rich food resources (e.g. nearer to the research station where
there is access to food waste). Alternatively, individuals that invest more energy in
foraging and less in vigilance, regardless of acclimation, might be easier to capture.
6.2.3. Prospects for stress physiology and human disturbance
This last section aims to relate the previous findings to stress physiology, and
provide expectations about the results on GC levels, with respect to the general
trend featured in the current literature.
The first set of results indicates that external parasites have no major effect on the
health of C. melanosterna. However, the costs of parasitism are often difficult to
detect because of other environmental and behavioural factors that might allow
infested individuals to compensate, as mentioned previously (Main and Bull, 2000).
However, parasitism is a known stressor to the host, thus the patterns in GC levels
can reveal whether the parasite has a negative effect. For example, it is often found
that animals with ectoparasites maintain a relatively high GC baseline level in order
to facilitate energy assimilation and cope with challenging parasitism (Raouf et al.,

40
2006). As a consequence, if parasitism has a negative effect, we would expect to
see a positive correlation between external parasite load and GC baseline level of
the studied individuals. Caution is however needed when interpreting these results,
as the relationship between circulating GC levels and parasite infestation is likely to
depend on the type of ectoparasite and level of infection (Eggert, Jodice and
O'Reilly, 2010).
Finally, the results suggest that anthropogenic disturbance does not affect body
condition, haematocrit level and external parasite load of C. melanosterna on Cayo
Menor. There is a possibility that individuals are able to acclimate to human
disturbance by adjusting energy investments in antipredatory behaviour, thereby
maintaining good health conditions. This behavioural adaptation might therefore
account for the fact that iguanas in regular contact with humans (at the RS) do not
exhibit a decrease in body condition, metabolic capacity and immune function,
compared to undisturbed iguanas (at the NPF).
These findings are consistent with studies investigating the effect of anthropogenic
disturbance on stress physiology of reptiles. Studies on tree lizards (Urosaurus
ornatus) indicate that individuals undergoing constant human disturbance in urban
areas have significantly lower baseline and stress-induced GC levels. However, the
amplitude of response is unchanged, indicating that these lizards can mount
appropriate response to environmental stressor (French, Fokidis and Moore, 2008).
Similarly, Romero and Wikelski (2002) investigated the effect of tourism on
Galápagos marine iguanas (Amblyrhynchus cristatus) and found that human
disturbed iguanas could mount appropriate GC responses upon capture, but lower
compared to undisturbed individuals. Again, this indicates that those individuals have
adapted to continuous human disturbance by lowering GC levels (baseline and acute
response). Nevertheless, it is unclear whether such acclimation can have negative
effects on long-term fitness (Romero and Wikelski, 2002).
If the spiny-tailed iguanas studied show similar acclimation, we would therefore
expect lower stress-induced GC response, and lower baseline level so that
appropriate stress response is still possible. It is unlikely that chronic stress can
account for the results obtained, as it is usually associated with a decrease in body
condition and an increase in parasitism resulting from a lack of immune response
(Busch and Hayward, 2009).

41
It is worth noting that results from previous studies show that GC levels can vary
depending on many factors, including predator pressure, infection status,
reproductive state, body condition, and environmental conditions (e.g. food
abundance, habitat change) (Busch and Hayward, 2009). Including these covariates
in the analysis is therefore crucial in order to identify differences in GC levels
resulting from anthropogenic disturbance.
6.3.Implications and future directions
The present results indicate that the current anthropogenic disturbance exerted on
the spiny-tailed iguana population of Cayo Menor is not causing health defects
(based on the health indicators used), even suggesting that iguanas have an
adaptive potential enabling them to acclimate to humans. This would confirm what
was previously suggested: spiny-tailed iguanas are robust to anthropogenic
disturbance (Suárez-Domínguez et al., 2011). On the other hand, down-regulation of
GC peak response to stressor could also mean that individuals become unable to
mount an appropriate stress response when needed (e.g. in the case of predation).
Acclimating to human disturbance would therefore be advantageous on the short-
term as it allows avoiding chronic stress and the costs associated with it, but is
potentially maladaptive on the long-term. To date, there appears to be no study
investigating to potential negative effects of acclimation on long-term fitness.
Regardless of acclimation, it is essential to quantify the effect that human
disturbance has on long-term fitness. In fact, Breuner, Patterson and Hahn (2008)
report that there is little empirical evidence directly linking acute GC response with
reproductive success and survival. Thus, C. melanosterna offers the opportunity to
explore the link between fitness (in terms of survival and reproductive output) and
anthropogenic disturbance, and the underlying physiological mechanisms.

42
7. ACKNOWLEDGEMENTS
I thank the Honduran Coral Reef Foundation (HCRF) and Operation Wallacea for
supporting the project. I would like to thank Dr Chad E Montgomery, Julius ‘Tony’
Frazier and Prof Roger Wotton for their help and supervision. I would also like to
thank Daniel Nicholson, Thaddaeus Cheng, Dora Sulej, and numerous volunteers for
the invaluable assistance they provided on the field. Finally, I am grateful to the
HCRF staff, the Operation Wallacea staff, the inhabitants of the Garifuna village East
End and the staff of Plantation Beach Resort on Cayo Grande for allowing us to work
in optimal conditions.

43
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Breuner, C.W., Patterson, S.H. & Hahn, T.P., 2008, In search of relationships
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48
ANNEXE 1
Table 1 Investigation of the variation in health condition due to sex, using ANOVA
tests (for BCI, HCT and tick load) and Chi-square test (for mite presence/absence).
Significant differences at the 0.05 confidence level are marked with *.
Sex class N Mean SE T (df) P value
Male 20 0.004674 0.01733
BCI
Female 41 0.000079 0.01210
0.217349
(59)
0.8287
Male 8 28.8750 1.6220
HCT
Female 26 31.2308 0.8997
-1.27009
(32)
0.2132
Male 19 8.57895 2.3987Tick
Number Female 42 2.14286 1.6133
2.226447
(59)
0.0298*
% Absence Sex class N % Absence ChiSquare P value
Male 19 73.68Mite
P/A
75.41
Female 42 76.19
0.044 0.8339
Table 2 Investigation of the variation in health condition due to life stage, using
ANOVA tests (for BCI, HCT and tick load) and Chi-square test (for mite
presence/absence).
Life stage N Mean SE F (df1, df2) P value
(Sub)Adult 48 0.00367 0.00939
Juvenile 16 -0.01078 0.01626BCI
Hatchling 83 -0.00301 0.00714
0.3363
(144, 2)
0.7150
(Sub)Adult 25 31.1200 0.9237
Juvenile 12 30.0000 1.3333HCT
Hatchling 3 28.3333 2.6666
0.6204
(37, 2)
0.5432
(Sub)Adult 48 5.12500 0.9928
Juvenile 16 0.50000 1.7195
Tick
Number
Hatchling 83 0.28916 0.7550
7.8840
(2, 144)
0.0006*
% Absence Life stage N % Absence ChiSquare P value
(Sub)Adult 48 75.00
Juvenile 16 75.00
Mite
P/A
83.67
Hatchling 83 90.36
6.231 0.0444*

49
ANNEXE 2
Table 1 Summary of the regression analyses (A, B, C) and t-tests (D, E, F) that were
used for paired comparisons of the different health indicators.
Indicators N Slope SE R2
T (df) P value
A
BCI
HCT
39 10.868723 8.883888 0.03888
1.22
(37)
0.2289
B
HCT
Ticks
40 -0.047011 0.335538 0.000516
-0.14
(38)
0.8893
C
BCI
Ticks
145 6.2045598 8.789086 0.003473
0.71
(143)
0.4814
2
1
Mites N Mean SE T (df) P value
Yes 12 29.6667 1.3259D HCT
No 28 30.9643 0.8680
-0.81883
(38)
0.4180
Yes 24 0.00030 0.01333
E BCI
No 121 -0.00155 0.00594
0.127076
(143)
0.8991
Yes 24 1.95833 1.4737
F
Tick
number No 123 1.87805 0.6510
0.049832
(145)
0.9603

50
ANNEXE 3
Table 1 Summary of the ANOVA and chi-square analyses used in order to compare
iguanas from the RS with those from the NPF, for each health indicator.
Indicator Site N Mean SE df T ratio P value
RS 122 -0.00195 0.00588BCI
NPF 25 -0.00033 0.01299
145 -0.11372 0.9096
RS 29 30.7241 0.8592
HCT
NPF 11 30.1818 1.3950
38 0.331019 0.7424
RS 122 2.24590 0.6497
Tick count
NPF 25 0.16000 1.4353
145 1.323937 0.1876
Site N N Absence % Absence ChiSquare P value
RS 122 102 83.61
NPF 25 21 84.00
0.002 0.9612
Total 147 123 83.67 - -
Table 2 Summary of the ANOVA and chi-square analyses used in order to
recaptured with newly captured iguanas, for each health indicator.
Indicator Capture N Mean SE df T ratio P value
New 26 -0.02916 0.01417BCI
Recapture 38 0.02005 0.01172
62 2.677103 0.0095*
New 17 30.5882 1.1076
HCT
Recapture 20 30.9000 1.0211
35
0.206949 0.8372
New 25 3.04000 2.1249
Ticks
Recapture 39 4.56410 1.7013
62
0.55991 0.5776
Capture N N Absence % Absence ChiSquare P value
New 25 20 80.00
Recapture 39 28 71.79
0.558 0.4550
Total 64 48 75.00 - -

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