PHYSIOLOGICAL ECOLOGY - ORIGINAL RESEARCH
Detrimental influence on performance of high temperatureincubation in a tropical reptile: is cooler better in the tropics?
Kris Bell • Simon Blomberg • Lin Schwarzkopf
Received: 5 April 2011 / Accepted: 21 June 2012 / Published online: 11 July 2012
� Springer-Verlag 2012
Abstract Global temperatures have risen over the last
century, and are forecast to continue rising. Ectotherms
may be particularly sensitive to changes in thermal
regimes, and tropical ectotherms are more likely than
temperate species to be influenced by changes in environ-
mental temperature, because they may have evolved nar-
row thermal tolerances. Keelback snakes (Tropidonophis
mairii) are tropical, oviparous reptiles. To quantify the
effects of temperature on the morphology and physiology
of hatchling keelbacks, clutches laid by wild-caught
females were split and incubated at three temperatures,
reflecting the average minimum, overall average and
average maximum temperatures recorded at our study site.
Upon hatching, the performance of neonates was examined
at all three incubation temperatures in a randomized order
over consecutive days. Hatchlings from the ‘hot’ treatment
had slower burst swim speeds and swam fewer laps than
hatchlings from the cooler incubation temperatures in all
three test temperatures, indicating a low thermal optimum
for incubation of this tropical species. There were no sig-
nificant interactions between test temperature and incuba-
tion temperature across performance variables, suggesting
phenotypic differences caused by incubation temperature
did not acclimate this species to post-hatching conditions.
Thus, keelback embryos appear evolutionarily adapted to
development at cooler temperatures (relative to what is
available in their habitat). The considerable reduction in
hatchling viability and performance associated with a
3.5 �C increase in incubation temperature, suggests climate
change may have significant population-level effects on
this species. However, the offspring of three mothers
exposed to the hottest incubation temperature were appar-
ently resilient to high temperature, suggesting that this
species may respond to selection imposed by thermal
regime.
Keywords Climate change � Developmental acclimation �Ectotherm � Performance � Phenotype
Introduction
Global temperatures have increased markedly since the
eighteenth century, and are predicted to rise at an
increasing rate over the next century (IPCC 2007). Tem-
perature has profound effects on the biological processes of
all animals (Birchard 2004). Ectotherms lack effective
physiological mechanisms to control body temperature,
and are therefore particularly sensitive to environmental
conditions (Bogert 1949). Because they have been exposed
to smaller variations in temperature over evolutionary time,
tropical ectotherms are likely to be more strongly influ-
enced by changes in environmental temperature than
temperate species (Huey et al. 2009; Sunday et al. 2010).
Embryos require appropriate levels of temperature, mois-
ture and gas exchange for successful development (Shine and
Thompson 2006). However, early stage embryos lack the
physiological and behavioural responses available to post-
hatching animals, suggesting that ectothermic organisms
may be most vulnerable to adverse thermal conditions at
Communicated by Mark Chappell.
K. Bell (&) � L. Schwarzkopf
School of Marine and Tropical Biology, James Cook University,
Townsville, QLD 4811, Australia
e-mail: [email protected]
S. Blomberg
School of Biological Sciences, The University of Queensland,
Brisbane, QLD 4072, Australia
123
Oecologia (2013) 171:83–91
DOI 10.1007/s00442-012-2409-6
early stages of development (Birchard 2004; Shine and
Thompson 2006). Thus, a warming climate has potentially
significant implications for embryogenesis in ectotherms.
Many oviparous ectotherms lay their eggs underground
(Booth 2006), presumably to provide protection from pre-
dation, and to avoid thermal and hydric extremes. The
thermal buffering properties of soil attenuate heat flux,
typically causing a complete absence of diel fluctuations in
temperature from depths of between 30 and 50 cm (Ack-
erman and Lott 2004). Despite little variation in tempera-
tures at even shallow soil depths, it is often implicitly
assumed that there are a wide range of thermal regimes
available, from which females can select suitable nest sites
(see Wilson 1998; Kolbe and Janzen 2002; Brown and
Shine 2004). Testing of this assumption is required if we
are to understand the potential susceptibility of oviparous
ectotherms to climate change.
The effects of climate change on ectotherms could
potentially be mitigated by acclimation. Several studies have
found that hatchlings prefer, or respond positively to,
ambient temperatures corresponding to temperatures expe-
rienced during development (Blouin-Demers et al. 2000;
Bronikowski 2000; Geister and Fischer 2007), suggesting
that acclimation may pre-adapt individuals to post-hatching
conditions (the Beneficial Acclimation Hypothesis; Leroi
et al. 1994). The majority of existing studies that examine the
plasticity of ectotherms to incubation regime test offspring at
only one temperature (see review by Deeming 2004). Such
an approach prevents the accurate assessment of differential
temperature-performance relationships (temperature reac-
tion norms), because it excludes the effects of phenotypic
acclimation (Seebacher 2005; Deere and Chown 2006).
Plasticity in the ecological and life history traits of
snakes make them good ‘model’ organisms in which to
examine available nest temperatures, offspring responses
to incubation temperature, and offspring acclimation to
incubation temperature (Shine and Bonnet 2000). How-
ever, many past studies do not base incubation temperature
treatments on ‘natural’ temperatures (Burger et al. 1987;
Gutzke and Packard 1987; Lin et al. 2005; Ji et al. 2007;
Du and Ji 2008). To determine the availability of suitable
nest sites to oviparous reptiles in a wetland ecosystem in
north-eastern Australia, we measured the magnitude of
thermal heterogeneity at different soil depths, by measuring
soil temperature profiles. We then used these temperature
profiles, together with current projections for climate over
the next century, as a basis to create appropriate incubation
treatments. We examined the morphological and pheno-
typic responses of hatchling keelback snakes (Tropidono-
phis mairii) to incubation temperatures. Evidence of
acclimation responses were examined by incubating eggs
from each clutch at three temperatures, then testing the
performance of all hatchlings at the three test temperatures.
Materials and methods
Study species
The freshwater keelback snake (Tropidonophis mairii,
Gray 1841) is a small, natricine colubrid that occurs in
wetland and woodland habitats throughout tropical Aus-
tralia (Cogger 1986) (Fig. 1). They are oviparous, and
natural nests occur in soil cracks and burrows at depths of
between 10 and 20 cm (Webb et al. 2001; Brown and Shine
2004).
Data collection
Soil temperature
To determine the range of temperatures available to keel-
backs in nest sites, 24 iButton� temperature data loggers
were deployed on 12th February 2010 at the Townsville
Town Common Conservation Park, northern Queensland
(19�12028.6900S, 146�44025.1700E). Locations for data log-
gers were based on randomly generated vectors (using
S-PLUS to generate values between -15 and 15, repre-
senting distances in metres for north/south and east/west
bearings) from catch locations of gravid female keelbacks.
At each location, we deployed iButtons� at depths that
reflected the range in which natural nests have been
observed (10–20 cm), as well as the temperature taken just
under the soil’s surface (2 cm depth) for comparison. The
iButtons� recorded temperatures hourly for a minimum of
2 weeks. The loggers were relocated on 3 March and 17
March 2010, thus providing temperature profiles for 24
locations adjacent to capture points, at a time of year when
eggs were likely to be in the soil. The proximity of vege-
tation, water and shade were recorded for each location.
Fig. 1 The keelback snake (Tropidonophis mairii)
84 Oecologia (2013) 171:83–91
123
Incubation
Thirty-six gravid female keelbacks were captured during
nightly surveys between 14 January and 25 February 2010
in the Townsville Town Common Conservation Park. The
adults were maintained in ventilated 37 9 30 9 20 cm
(L 9 W 9 H) plastic bins, provided with water ad libitum,
a 650-ml hide containing a substrate of vermiculite and
water (1:1 by mass), and a ‘heat mat’ for thermoregulation,
and allowed to lay eggs. Containers were checked twice
daily for eggs. Immediately following discovery, eggs were
weighed and measured (maximum length and width in mm
using callipers) before being placed in separate 300-ml
plastic tubs containing 100 ml of vermiculite substrate
moistened with 10 ml of water, and covered in plastic cling
film to prevent excessive water loss. Eggs were then ran-
domly assigned, in a split-clutch design, to one of three
temperature treatments (24.9 �C ± 0.7, 26.6 �C ± 0.9,
and 30.1 �C ± 0.5) until they hatched. Temperatures
within each treatment fluctuated by the same magnitude as
those recorded at the study site. The coolest temperature
treatment (hereafter referred to as ‘cold’) reflected the
average minimum daily temperature, at a 20-cm depth, of
the coldest iButton location (24.8 �C ± 0.8). The inter-
mediate temperature incubation treatment (‘mid’) reflected
the average daily temperature, at a depth of 20 cm, across
all iButton locations (26.2 �C ± 1.0). The ‘hot’ incubation
treatment reflected the average maximum daily tempera-
ture, at 20-cm depth, of the warmest iButton location
(29.7 �C ± 0.7). The hot treatment also reflected the pre-
dicted average temperature at a depth of 20 cm (or current
‘mid’ treatment) by 2090–2099, given current best esti-
mates of climate change (IPCC 2007). Humidity in all
three rooms was held constant at approximately 80 %.
Eggs were checked daily and were re-arranged within each
room every 2 days to avoid position effects. Following
oviposition, females were released at their point of capture.
Upon hatching, neonates were weighed, measured
[snout-vent length (SVL), tail length, head width and
length] to the nearest millimetre using callipers, sexed by
eversion of hemipenes and checked for the presence of
ventral scale asymmetries, which can indicate develop-
mental instability (Shine et al. 2005; Lowenborg et al.
2010). Once measured, neonates were placed in ventilated
650-ml containers, provided with water ad libitum and
randomly assigned to one of the three temperature treat-
ments. Following an acclimation period of 24 h, hatchlings
were tested (see below) before being transferred to the next
(randomly assigned) treatment room and allowed to accli-
mate for a further 24 h, so that each snake was tested over
3 consecutive days in each temperature treatment. Once
testing was complete, the neonates were held in conditions
similar to those described for gravid females above, until a
first slough had been completed, and then they were
released at the point of capture of the mother.
The effects of incubation treatment on hatchlings were
assessed over a range of performance measures. Locomotor
ability was assessed using video-recorded swim trials.
Keelbacks live near water and often swim to escape pre-
dation and to forage for food (Webb et al. 2001). Snakes
were placed in a circular swim chamber (outer diameter,
120 cm; inner diameter, 80 cm) filled with water to a depth
of 4 cm. The water used in swim trials was left to stand in
each treatment room for a minimum of 24 h prior to test-
ing, and water temperatures were measured immediately
prior to swim trials. The hatchlings were encouraged to
swim by gently tapping their tails with a test tube brush.
Each snake was allowed to swim for 5 min, and the number
of times they were tapped, the number of laps completed
and the time of the first escape attempt (whereby hatchlings
attempted to crawl up the sides of the chamber) were
recorded. The fastest time taken by each hatchling to cover
a 1/8th section (47 cm) of the circuit was measured by
examination of the videos, and used as a measure of burst
speed.
Data analysis
Incubation
There were significant correlations between SVL, tail
length, and head length/width of hatchlings, so a principal
components analysis (PCA) was conducted on the covari-
ance matrix to reduce the dimensionality of morphological
traits. PC1 and PC2 were used as dependent variables in
unbalanced, mixed-effects ANOVAs, with clutch-of-origin
as a random, block effect and incubation treatment as a
fixed effect. Body mass scaled differently from length and
width measurements, and was therefore not included in the
PCA. Variables that violated the assumptions of ANOVA
were log-transformed prior to analysis.
We used linear mixed-effects models (LMM) and gen-
eralised linear mixed-effects models (GLMM) to analyse
snake performance. Clutch of origin and snake ID (nested
within clutch) were included as random factors. Incubation
treatment and test temperature treatment (and their inter-
action) were included as fixed effects, and PC1 was
included as a covariate. For the number of times tapped, a
Poisson GLMM with a square-root link function was used.
LMMs were used for burst speed and number of laps. Plots
of the (weighted) residuals versus the fitted values were
examined to check for heteroskedasticity, and normal
quantile–quantile plots were examined to check for
departures from normality of the residuals. Analyses were
performed in R (R Development Core Team 2012), using
the lme4 package (Bates et al. 2011). p values for the fixed
Oecologia (2013) 171:83–91 85
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effects in LMMs were calculated using Markov Chain
Monte Carlo (MCMC) methods, using the languageR
package for R (Baayen 2011). Significance of the fixed
effects in GLMM was tested using likelihood ratio Chi-
square tests. Post hoc comparisons of fixed effects were
conducted using the package multcomp for R (Hothorn
et al. 2008). Significance of random effects were assessed
using likelihood ratio Chi-square tests.
Results
Soil temperature
The average temperature recorded across all 24 sites and
depths between February and May 2010 was 26.2 �C (±2.3
SE). There was a significant difference in temperature (t test,
t(12989) = 14.36, p \ 0.001) and variance (F test, F(10318,
10788) = 7.29, p \ 0.001) among sites exposed to the sun and
those in the shade. Shaded sites (26.1 �C ± 1.1) were cooler
and exhibited less variation in temperature than sites exposed
to the sun (26.5 �C ± 3.1). Temperature and variation dif-
fered significantly among 2, 10 and 20 cm depths (means:
Kruskall–Wallis, v2ð2Þ ¼ 158:7, p \ 0.001; Variances:
Levene’s Homogeneity of Variance test(2,35351) = 2,112.3,
p \ 0.001). Temperatures 2 cm underground (26.5 �C ±
3.5) were warmer and varied more than at 10 cm
(25.9 �C ± 1.3) and 20 cm (26.2 �C ± 1.0) (Fig. 2).
Incubation
Egg weight, width and length were not significantly different
among incubation treatments [egg weight, mean = 3.0 g
(cold, mid, hot), F(2,390) = 0.02, p = 0.98; egg width,
mean = 25.7 mm (cold), 25.8 mm (mid), 26.1 mm (hot),
F(2,390) = 0.37, p = 0.70; egg length, mean = 14.0 mm
(cold), 14.1 mm (mid), 14.0 mm (hot), F(2,390) = 0.71,
p = 0.49], so the random allocation of eggs among treat-
ments was effective. Across all treatments, the number of
female hatchlings (103) did not differ significantly from
males (112) (Binomial test, n = 215, p = 0.59). Also,
there were no significant differences in sex ratios among
incubation treatments (Pearson, v2ð2Þ ¼ 2:80, p = 0.25).
After hatching, hot-incubated snakes took significantly
longer (11.4 days ± 0.9) to slough than snakes from
mid (7.2 days ± 0.3) or cold (7.5 days ± 0.3) incubation
treatments (Kruskall–Wallis, v2ð2Þ ¼ 6:65, p = 0.036;
Table 1).
Hatching success
Incubation time was inversely related to incubation tem-
perature, with neonates from the hottest incubation tem-
perature hatching earliest (41.4 ± 1.4 days), followed by
mid (51.0 ± 2.3 days) then cold (60.7 ± 2.2 days)
(ANOVA, F(2,189) = 3,253.6 p \ 0.001). Hatching success
decreased significantly with increasing incubation temper-
ature (v2ð2Þ ¼ 107:8, p \ 0.001), with a hatching success
rate of 79.2 and 72.9 % in cold and mid incubation tem-
peratures respectively, dropping to a success rate of 21.4 %
in the hottest temperature treatment. The probability of a
neonate hatching successfully from the hottest incubation
temperature was not distributed randomly among mothers,
with significant clustering in hatching success amongst
clutches (three mothers had an average hatching success of
85 %, compared to a hatching success of 14 % on average
for all other mothers, Poisson, v2ð4Þ ¼ 22:5, p \ 0.001).
Clutches in the hottest incubation temperature were more
likely to have many successful hatchings or none at all, and
significantly less likely to produce intermediate numbers of
viable offspring than would be expected if hatching success
was random (Fig. 3).
Morphology
The number and proportion of ventral scale asymmetries
differed significantly among incubation temperatures
(v2ð2Þ ¼ 110:6, p \ 0.001), with the hottest treatment show-
ing the highest prevalence (85.7 %, 24/28), followed by cold
(6.9 %, 7/101), then mid (6.4 %, 6/94). Similarly, the
occurrence of other developmental abnormalities (typically
post-cloacal kinking of the spine) was significantly different
among incubation treatments (v2ð2Þ ¼ 111:1, p \ 0.001).
71.4 % (20/28) of hot incubated snakes displayed someFig. 2 Average daily soil temperatures at the study site at different
soil depths
86 Oecologia (2013) 171:83–91
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visible form of developmental abnormality, compared to just
3.2 % (3/94) and 3.0 % (3/101) of mid- and cold incubated
snakes respectively.
PC1 (56.2 %) and PC2 (19.6 %) explained 75.8 % of
the total variance among morphological variables (Fig. 4).
The morphological variables with the highest loadings on
PC1 were tail length (0.81) and SVL (0.79). Head width
(0.79) had a considerably larger loading on PC2 than all
other variables, thus PC1 and PC2 appear to reflect body
‘size’ and body ‘shape’ respectively. Body size (PC1)
differed significantly among incubation treatments, after
correcting for maternal effects (ANOVA, F(2,189) = 34.9,
p \ 0.001) (Fig. 3). Hatchlings from the hot incubation
treatment were significantly smaller than both cold and mid
incubation treatments, though cold and mid incubation
temperatures did not differ from one another. Body shape
(PC2), particularly head width, was also significantly dif-
ferent among the three treatments (ANOVA, F(2,189) =
10.9, p \ 0.001). Cold and hot incubated hatchlings had a
slimmer build than hatchlings from the mid incubation
temperature.
Hatchlings from the hot incubation treatment (2.46 g ±
0.30) weighed significantly less than hatchlings from the
mid (2.68 g ± 0.44) and cold treatments (2.75 g ± 0.44)
(ANOVA, F(2,188) = 9.0, p \ 0.001).
Performance
Burst speed was significantly influenced by both incubation
temperature (LMM, F(2,x) = 82.75, MCMC p \ 0.0001
and the temperature at which trials were conducted (LMM,
F(2,x) = 13.01, MCMC p = 0.0004) (Fig. 5a). The inter-
action between incubation temperature and trial tempera-
ture was not significant (LMM, F(4,x) = 0.58, MCMC
p = 0.7742), after correcting for body size, maternal
effects and ID effects.
Body size was significantly positively related to burst
speed (slope = 1.756, t(x) = 2.852, MCMC p = 0.0011).
The proportion of variance associated with maternal effects
was only 7 %, and this was not significantly different from
zero (LMM, LR v2ð1Þ ¼ 2:6915, p = 0.1009). The propor-
tion of variance explained by individual snake ID was
higher and significant at 28 % (LMM, LR v2ð1Þ ¼ 15:676,
p \ 0.0001). After averaging over body size and the
incubation 9 test interaction, the hot incubation treatment
had significantly lower burst speeds than either the cold
treatment (Tukey HSD test, z = 6.469, p \ 0.0001) or the
mid treatment (Tukey HSD, z = 5.187, p \ 0.0001). The
Cold treatment was not significantly different from the mid
treatment (Tukey test, z = -1.234, p = 0.433).
Table 1 Effects of incubation treatment on hatching success, mor-
phology, behaviour and performance in hatchling keelback snakes
(Tropidonophis mairii)
Variable Incubation temperature, mean (SD)
Cold
(n = 106)
Mid
(n = 94)
Hot
(n = 27)
Sex ratio (M:F) 44:57 46:44 9:15
Time to first shed
(days)
7.5 (0.3) 7.2 (0.3) 11.4 (0.9)*
Hatchability
Incubation time
(days)
60.7 (2.2)* 51.0 (2.3)* 41.4 (1.4)*
Hatching success
(%)
79.2 72.9 21.4*
Residual egg
weight (g)
0.69 (0.23) 0.61 (0.28) 0.75 (0.39)
Morphology
Mass (g) 2.7 (0.4) 2.7 (0.4) 2.5 (0.3)*
SVL (cm) 15.6 (1.0) 15.3 (0.8) 14.7 (0.9)*
Tail length (cm) 4.4 (0.5) 4.4 (0.5) 4.0 (0.4)*
Head width (mm) 4.6 (0.1)* 4.6 (0.2) 4.5 (0.2)
Head length (mm) 9.7 (0.3) 9.6 (0.3) 9.4 (0.4)
Ventral scale
asymmetries (%)
6.9 6.4 85.7*
Spinal deformities
(%)
3.0 3.2 71.4*
Performance
Burst speed (cm/s) 22.0 (7.1) 19.9 (6.4) 8.0 (6.1)*
Number laps 5.4 (0.1) 5.45 (0.1) 2.3 (0.3)*
Number taps 12.0 (0.6) 13.7 (0.6) 6.9 (1.2)*
* Statistically different subgroup
Fig. 3 Frequencies of observed and expected hatchling numbers
from clutches incubated in the warmest temperature treatment. Note
that many more clutches had no hatchlings than expected in the hot
treatment
Oecologia (2013) 171:83–91 87
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An additional measure of swim speed, the number of
laps completed in 5 min, also showed significant differ-
ences with incubation temperature (F(2,x) = 113.94,
MCMC p \ 0.0001) and performance test temperature
(F(2,x) = 8.211, MCMC p \ 0.0001), with no significant
interaction between the two factors (F(4,x) = 1.952, MCMC
p = 0.16159). See Fig. 5b. Again, there was a significant
effect of body size, with larger individuals swimming further
(slope = 0.3180, tx = 3.543, MCMC p \ 0.0001). Mater-
nal and individual components of variance were 7.6 %
(LLMM, LR v2ð1Þ ¼ 8:524, p = 0.0035), and 16.6 %
(LLMM, LR v2ð1Þ ¼ 17:983, p \ 0.0001), respectively. Post
hoc comparisons of means (averaged over the fixed factor
interaction and the body size covariate) revealed that the hot
incubated animals swam fewer laps than either the cold
(Tukey HSD, z = -11.941, p \ 0.0001) or mid temperature
(Tukey HSD, z = 12.301, p \ 0.0001) incubated animals,
but that the cold and mid temperature incubation treatments
were not significantly different (Tukey HSD, z = 0.452,
p = 0.89). Post hoc analysis of the performance temperature
treatments showed that there was a significant difference
between hot and cold running temperatures (Tukey HSD,
z = 2.919, p = 0.0098), but not between hot and mid
(Tukey HSD, z = -1.967, p = 0.121) or between cold and
mid (Tukey HSD, z = 0.951, p = 0.608), indicating that the
effect of performance temperature on the number of laps
swum in 5 min is less than the effect of incubation temper-
ature on this response variable.
Willingness to swim, as measured by the number of taps
required during testing (Brown and Shine 2002), showed a
significant interaction between incubation temperature and
performance temperature (LR v2ð4Þ ¼ 11:586, p = 0.0207;
see Fig. 5c). Post hoc tests for the incubation temperature
effect (averaged over the interaction term and body size
covariate) revealed a significant difference between both
hot and cold incubation treatments (Tukey HSD, z =
-4.017 p \ 0.001) and hot and mid incubation treatments
(Tukey HSD, z = -5.120, p \ 0.001) but not mid and cold
incubation treatments (Tukey HSD, z = 1.757, p = 0.177).
Thus, hatchlings from the hot incubation treatment were
more willing to swim than hatchlings from both mid and
cold incubation treatments. An equivalent analysis of the
performance treatments revealed no significant difference
between the mid and cold treatments (Tukey HSD,
z = 0.415, p = 0.91) but differences between the hot and
cold treatments (Tukey HSD, z = 6.614, p \ 0.0001) and
between hot and mid treatments (Tukey HSD, z = 6.198,
p \ 0.0001). Hence, the hot performance temperature
required less taps than either the mid or cold performance
temperature treatments.
Discussion
Clearly, keelback snakes show high levels of phenotypic
plasticity in response to thermal regime; incubation
Fig. 4 Principal components
analysis plotting body ‘size’
(PC1) and body ‘shape’ (PC2)
in hatchling keelback snakes
from hot (circles), mid
(squares) and cold (crosses)
incubation treatments. Darkbold markers indicate
centroids ± standard errors
88 Oecologia (2013) 171:83–91
123
temperature significantly affected 12 of the 15 traits
examined. The temperature at which hatchlings were
incubated had a stronger influence on performance than did
test temperature across all performance measures, includ-
ing locomotor ability and willingness to swim.
Performance was ‘‘best’’, i.e., offspring had fewer abnor-
malities, were larger, and could swim faster and for longer,
if they had experienced cold and mid temperatures as
embryos, suggesting that development is evolutionarily
adapted to occur at these temperatures. Apparent adapta-
tion to cooler temperatures is consistent with a surprisingly
low optimal incubation temperature in this tropical reptile
(Brown and Shine 2006). The only characteristic that was
not ‘‘best’’, or of apparently higher fitness, in offspring
incubated in the two cooler treatments was willingness to
swim. Smaller, slower offspring from warmer temperatures
swam more readily when coaxed compared to offspring
from other treatments. This may be due to covariation
among physiological traits, such that high speed is a trade-
off against higher endurance (or willingness to swim)
(Vanhooydonck et al. 2001). However, many studies dis-
pute the existence of significant trade-offs between speed
and endurance (Huey et al. 1990; Secor et al. 1992, 1995;
Brodie and Garland 1993; Goodman et al. 2007). Alter-
natively, perhaps increased willingness to swim is a
behavioural adaptation that mitigates smaller size and
slower swimming speeds. For all other characters, both
behavioural and morphological, cooler temperatures
appeared to produce ‘fitter’ offspring.
In our study, individuals exposed to cooler incubation
temperatures (similar to cooler ranges of temperatures
available in the wild) showed higher performance in all
incubation temperatures. The lack of interaction between
incubation temperatures and test temperatures indicates
that developmental acclimation did not pre-adapt individ-
uals to post-hatching environments. Cooler incubation
temperatures are often better in terms of hatchling fitness in
ectotherms (Partridge et al. 1995; Atkinson and Sibly 1997;
Shine 1999; Stillwell and Fox 2005; Brown and Shine
2006). However, our study does not really support the
‘cooler is better’ hypothesis of thermal acclimation,
because we tested only the range of temperatures actually
available to female snakes in the wild. Thus, we did not
demonstrate that colder temperatures in general produce
fitter hatchlings, rather we showed that, within the range of
temperatures available to females, cooler temperatures
were better for successful embryonic development. These
results are, therefore, also consistent with the local adap-
tation hypothesis of thermal acclimation (Caley and
Schwarzkopf 2004), because females apparently select cool
nest sites, at least in other regions of Australia (Brown and
Shine 2006).
Preference for relatively cool nest sites may increase the
sensitivity of T. mairii to global increases in environmental
temperatures. The 3.4 �C difference in average soil tem-
peratures between the hottest and coldest sites (at a depth
of 20 cm) suggests that the availability of suitable nest sites
may be limited, even in relatively complex environments.
Fig. 5 Incubation treatment and test temperature effects on the
performance of hatchling keelback snakes. a, b Measures of
locomotor speed, c willingness to swim (note: more taps indicate
snakes were less willing). The x axis represents the temperature at
which snakes were tested. The y axis represents the means of the
dependent variables. Hatchlings incubated at the cold temperature
treatment are indicated by the dotted black line, mid temperature by
the grey line and hot by the solid black line. Vertical bars standard
errors
Oecologia (2013) 171:83–91 89
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Global average surface temperatures are forecast to rise by
up to 4 �C by 2100 (IPCC 2007). Should surface temper-
atures increase as predicted, then not one of the 24 sites at
which soil temperature profiles were recorded would pro-
vide optimal conditions for development of T. mairii.
However, despite the apparently small variation in soil
temperature, gravid keelbacks may still be able to locate
cool nest sites, and therefore they may be able to com-
pensate behaviourally for the influences of higher envi-
ronmental temperatures. Reptiles may adjust behaviour to
select thermally appropriate sites that enhance their fitness,
and expand or alter their geographic range (Huey et al.
2003). For example, grass snakes (Natrix natrix) have
extended their range in Europe into high latitudes by taking
behavioural advantage of heat sources such as manure piles
(Lowenburg 2010). Alternatively, compensation may occur
through temporal shifts in the breeding season, such as
laying eggs earlier in the spring. However, the importance
of moisture to the viability of eggs, and the correlation of
moisture levels with season, may restrict the extent to
which keelbacks could alter the timing of breeding (Brown
and Shine 2004). Given the detrimental impacts on
hatchling viability of relatively minor differences in incu-
bation temperature, further investigation into the presence
of cooler microclimates may be critical in predicting how
this species responds to climate change.
Another way that snake populations may respond to
global temperature increases is evolutionarily (Bradshaw
and Holzapfel 2006; Kearney et al. 2009). Although all the
clutches were highly sensitive to incubation temperature,
inconsistent hatching success among dams in the hot
incubation treatment suggests there may be genetic dif-
ferences in the thermal tolerance of individuals within
populations. The presence of apparently ‘heat tolerant’
dams suggests this population may be resilient to some of
the impacts of climate change. Identifying the genetic basis
for these differences may provide insight into the extent of
resilience in other populations of keelbacks, and poten-
tially, other species of snake.
While we have described the likely short-term conse-
quences of increased incubation temperature, little is known
about how specific phenotypic changes lead to differences in
post-hatching survival and, ultimately, fitness. Survival can
be negatively, positively or uncorrelated to incubation tem-
peratures within turtles, lizards and crocodiles (Deeming
2004). Limited evidence from mark–recapture studies on
keelbacks suggests that large sizes at hatching enhance sur-
vival (Brown and Shine 2004). Despite a lack of information
on long-term compensation effects, and the relationship
between specific traits and survivorship, a relatively small
increase in incubation temperature can considerably reduce
viability and performance of hatchling keelbacks. Hot-
incubated hatchlings had a lower hatching success rate, were
smaller and had more developmental abnormalities than
hatchlings from the mid and cold incubation treatments.
Climate change is predicted to cause local reptile extinctions
of 39 % worldwide by 2080 (Sinervo et al. 2010). Further
studies examining the links between phenotype and fitness,
the availability of suitable nest sites and the mechanisms
behind the high variability in phenotypic response to incu-
bation temperature, are critical to understanding how ovip-
arous reptiles will respond to a changing climate.
Acknowledgments We thank Teresea Lambrick, Gus McNab, Ross
Gottlieb, Joel Voiselle, John Llewelyn and Matthew Crowther for
their assistance in the field and laboratory. The work was carried out
under a permit from the Environmental Protection Agency and
Queensland Parks and Wildlife Service (WITK06346309), and James
Cook University Animal Ethics Committee (A1469). This research
was supported financially by a grant from the National Climate
Change Adaptation Research Facility.
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