DISCUSSION:
Patterns of population-level variation in embryonic responses to
maternal provisioning and environmental factors have the potential to
inform how the developmental environment contributes to evolutionary
change. We observed that, generally, the influence of maternal
provisioning on hatchling traits did not vary across populations;
however, incubation temperature exerted population-specific effects on
both morphological and metabolic traits. This may be explained by a
constrained relationship between egg mass and hatch mass (Deeming and
Birchard, 2007), which is expected to be maximized as hatchling mass is
often an important component of survival and fitness (Ronget et al.,
2018; Stearns, 2000). Thus, selection instead tends to act on aspects of
maternal allocation, such as egg size and number, to best match
population-specific conditions (Angilletta et al., 2004; Sinervo, 1990).
On the other hand, responses to incubation temperature may be in part
the result of differences in natural nest temperatures across
populations, which has been shown in several species (Du et al., 2019),
including the alligator (Bock et al., 2020). Such differences likely
select for embryonic responses to temperature that match
population-specific conditions. Our results suggest that plastic
responses to incubation temperature, but not maternal provisions, are a
source of interpopulation trait variation and may be more likely to be
modified by selection.
The four populations examined in this study encompassed a large
proportion of the alligator’s latitudinal range, with two populations
from the northern extent and two populations from the southern extent.
While not statistically significant, we observed a trend for smaller egg
masses at the northern populations relative to the southern populations.
In crocodilians, egg mass scales with maternal body size (Larriera et
al., 2004), and differences in maternal size might underlie population
differences observed here. In mammals, animals from high latitudes tend
to be larger than those from low latitudes in a pattern known as
Bergmann’s rule (Blackburn et al., 1999), and while this seems to hold
in turtles and birds, it does not in other reptiles (Ashton, 2002;
Ashton and Feldman, 2003) and has not been examined in crocodilians.
Nonetheless, larger egg sizes at southern populations might suggest the
opposite pattern. Interestingly, however, allometric relationships
between maternal size and egg mass can be altered by environmental
conditions, such as salinity (Murray et al., 2013). Whether differences
in egg size observed here are the result of differences in maternal size
across populations (maximum size or age at reproduction) or
population-specific allometric relationships is unknown and an
interesting area of future research.
While we expected to find responses to incubation temperature consistent
with latitudinal differences between our population pairs, only a few
traits showed such patterns. Namely, incubation duration was more
strongly influenced by incubation temperature at the northern
populations relative to the southern populations. Latitudinal
differences in incubation duration have been shown in several species
and generally follow one of two patterns: co-gradient variation, in
which cooler populations development more slowly relative to warmer
populations and counter-gradient variation, in which cooler populations
development more quickly than warmer populations (Conover and Schultz,
1995; Pettersen, 2020). Our results show embryos from northern
populations develop slightly slower at cooler temperatures and faster at
warmer temperatures compared to southern populations. While differences
within temperatures were not significant, they followed patterns of both
co-gradient variation (at 29.5°C) and counter-gradient variation (at
33.5°C). Similar results have been shown in Asian pond turtles
(Mauremys mutica ; Zhao et al., 2015) and may suggest that the
mechanisms responsible for variation in incubation duration across
populations may be temperature specific. On the other hand, increased
plasticity in developmental rate at the northern populations may be
driven by more variable thermal environments, which have been associated
with increased levels of physiological plasticity (Seebacher et al.,
2015). Additional experiments incorporating more incubation treatments
and populations are needed to more completely discern how the
relationship between temperature and developmental rate differs across
populations as well as the underlying mechanisms responsible.
Interestingly, we also observed that southern populations tended to
allocate more resources towards fat body mass than northern populations
at both incubation temperatures. The role of the fat body in alligators
is not known, and further work examining its function, including how fat
body size/mass early in life might impact survival and later life
fitness, is needed to more fully appreciate the potential consequences
of these patterns.
Apart from latitudinal trends, there were several differences in the
influence of incubation temperature between population pairs,
specifically between YK and other populations. In alligators, animals at
33.5°C are generally larger in mass than those at 29.5°C (Bock et al.,
2021), which was upheld across all populations. However, at YK, the
reduction of hatchling mass at 29.5°C appeared particularly pronounced
and appeared to drive additional phenotypic differences. Hatchling mass
relative to egg mass reflects the efficiency by which maternal resources
are converted into hatchling tissue and is likely a product of the
energetic cost of embryonic development (Pettersen et al., 2019). The
reduction in mass at YK at 29.5°C relative to the other sites suggests
that development at 29.5°C at YK was particularly inefficient.
Interestingly, however, animals at 29.5°C at YK tended to have residual
yolk reserves that were larger or equivalent to other populations after
controlling for mass. This may suggest that alligator embryos
preferentially allocate resources towards residual yolk mass at the cost
of reduction in overall size under sub-optimal conditions, which has
also been shown in other reptiles (Murphy et al., 2020; Radder et al.,
2004).
The lack of latitudinal trends in several of the morphological and
metabolic traits examined here suggests that latitude may not be the
best or only microclimatic proxy within which to understand variation in
responses to the developmental environment, particularly incubation
temperature. A similar lack of latitudinal patterns in response to
incubation temperature was shown across several populations of painted
turtles (Chrysemys picta ), another TSD species (Bodensteiner et
al., 2019). These results may be driven by too broad a resolution of the
relationship between temperature and latitude, making it impossible to
discern subtle population differences, or other population-specific
microclimatic variables that put selective pressure on thermal reaction
norms. For instance, of the four populations examined, YK is the only
coastal site, surrounded by brackish water, which may impose unique
selective pressures on embryos and breeding females, resulting in
differences in response to incubation temperature (e.g., Hudak and
Dybdahl 2023). Additionally, other maternal effects, such as yolk
composition and deposition of hormones and anthropogenic contaminants,
may, in addition to temperature, influence phenotype (Bae et al., 2021;
Du et al., 2010b; Groothuis et al., 2005), but were not considered here.
Further, since our design focused on incubation temperatures that
produce 100% males or females, population variation in response to
incubation temperature may have been driven by sex differences that
would not be explained by latitude. While previous work has shown that
phenotypic differences between incubation temperatures are the result of
temperature and not sex (Bock et al., 2023, preprint), whether sex
differences exist across populations irrespective of temperature is not
known. Future work examining the latter and the role of additional
aspects of the developmental environment as potential drivers of
variable responses to temperature across populations and the consistency
of such effects across years will be particularly informative.