Different optimal offspring sizes for sons versus daughters may favor the evolution of temperature-dependent sex determination in viviparous lizards

Temperature-dependent sex determination (TSD) has evolved independently in at least two lineages of viviparous Australian scincid lizards, but its adaptive significance remains unclear. We studied a montane lizard species (Eulamprus heatwolei) with TSD. Our data suggest that mothers can modify the body sizes of their offspring by selecting specific thermal regimes during pregnancy (mothers with higher and more stable temperatures produced smaller offspring), but cannot influence sons versus daughters differentially in this way. A field mark-recapture study shows that optimal offspring size differs between the sexes: larger body size at birth enhanced the survival of sons but reduced the survival of daughters. Thus, a pregnant female can optimize the fitness of either her sons or her daughters (via yolk allocation and thermoregulation), but cannot simultaneously optimize both. One evolutionary solution to reduce this fitness cost is to modify the sex-determining mechanism so that a single litter consists entirely of either sons or daughters; TSD provides such a mechanism. Previous work has implicated a sex difference in optimal offspring size as a selective force for TSD in turtles. Hence, opposing fitness determinants of sons and daughters may have favored evolutionary transitions from genetic sex determination to TSD in both oviparous turtles and viviparous lizards.     

The adaptive significance of temperature-dependent sex determination: experimental tests with a short-lived lizard

Abstract Why is the sex of many reptiles determined by the temperatures that these animals experience during embryogenesis, rather than by their genes? The Charnov‐Bull model suggests that temperature‐dependent sex determination (TSD) can enhance maternal fitness relative to genotypic sex determination (GSD) if offspring traits affect fitness differently for sons versus daughters and nest temperatures either determine or predict those offspring traits. Although potential pathways for such effects have attracted much speculation, empirical tests largely have been precluded by logistical constraints (i.e., long life spans and late maturation of most TSD reptiles). We experimentally tested four differential fitness models within the Charnov‐Bull framework, using a short‐lived, early‐maturing Australian lizard (Amphibolurus muricatus) with TSD. Eggs from wild‐caught females were incubated at a range of thermal regimes, and the resultant hatchlings raised in large outdoor enclosures. We applied an aromatase inhibitor to half the eggs to override thermal effects on sex determination, thus decoupling sex and incubation temperature. Based on relationships between incubation temperatures, hatching dates, morphology, growth, and survival of hatchlings in their first season, we were able to reject three of the four differential fitness models. First, matching offspring sex to egg size was not plausible because the relationship between egg (offspring) size and fitness was similar in the two sexes. Second, sex differences in optimal incubation temperatures were not evident, because (1) although incubation temperature influenced offspring phenotypes and growth, it did so in similar ways in sons versus daughters, and (2) the relationship between phenotypic traits and fitness was similar in the two sexes, at least during preadult life. We were unable to reject a fourth model, in which TSD enhances offspring fitness by generating seasonal shifts in offspring sex ratio: that is, TSD allows overproduction of daughters (the sex likely to benefit most from early hatching) early in the nesting season. In keeping with this model, hatching early in the season massively enhanced body size at the beginning of the first winter, albeit with a significant decline in probability of survival. Thus, the timing of hatching is likely to influence reproductive success in this short‐lived, early maturing species; and this effect may well differ between the sexes.     

Reproducing lizards modify sex allocation in response to operational sex ratios

Sex-allocation theory suggests that selection may favour maternal skewing of offspring sex ratios if the fitness return from producing a son differs from that for producing a daughter. The operational sex ratio (OSR) may provide information about this potential fitness differential. Previous studies have reached conflicting conclusions about whether or not OSR influences sex allocation in viviparous lizards. Our experimental trials with oviparous lizards (Amphibolurus muricatus) showed that OSR influenced offspring sex ratios, but in a direction opposite to that predicted by theory: females kept in male-biased enclosures overproduced sons rather than daughters (i.e. overproduced the more abundant sex). This response may enhance fitness if local OSRs predict survival probabilities of offspring of each sex, rather than the intensity of sexual competition.     

Reproductive ecology of the jacky dragon (Amphibolurus muricatus): an agamid lizard with temperature‐dependent sex determination

Abstract The jacky dragon, Amphibolurus muricatus (White, ex Shaw 1790) is a medium sized agamid lizard from the southeast of Australia. Laboratory incubation trials show that this species possesses temperature‐dependent sex determination. Both high and low incubation temperatures produced all female offspring, while varying proportions of males hatched at intermediate temperatures. Females may lay several clutches containing from three to nine eggs during the spring and summer. We report the first field nest temperature recordings for a squamate reptile with temperature‐dependent sex determination. Hatchling sex is determined by nest temperatures that are due to the combination of daily and seasonal weather conditions, together with maternal nest site selection. Over the prolonged egg‐laying season, mean nest temperatures steadily increase. This suggests that hatchling sex is best predicted by the date of egg laying, and that sex ratios from field nests will vary over the course of the breeding season. Lizards hatching from eggs laid in the spring (October) experience a longer growing season and should reach a larger body size by the beginning of their first reproductive season, compared to lizards from eggs laid in late summer (February). Adult male A. muricatus attain a greater maximum body size and have relatively larger heads than females, possibly as a consequence of sexual selection due to male‐male competition for territories and mates. If reproductive success in males increases with larger body size, then early hatching males may obtain a greater fitness benefit as adults, compared to males that hatch in late summer. We hypothesize that early season nests should produce male‐biased sex ratios, and that this provides an adaptive explanation for temperature‐dependent sex determination in A. muricatus.     

Seasonal sex ratios and the evolution of temperature-dependent sex determination in oviparous lizards

Although the adaptive significance of temperature-dependent sex determination (TSD) remains a puzzle, recent models implicate a seasonal bias in offspring sex production that translates into sex-specific fitness benefits later in life. Sex-specific emergence has been linked to fitness gains in some fish, birds and reptiles, but field data supporting the occurrence of a seasonal pattern of sex ratios in oviparous lizards are lacking. We tested the hypothesis that patterns of nest site selection and seasonal temperature changes combine to inhibit the materialization of sex-biased hatching times in a population of water dragons (Intellagama lesueurii). As predicted, a seasonal increase in air and nest temperatures resulted in a sex bias by nesting date; male-producing clutches were laid 17.8 days sooner than female-producing clutches, on average. However, the seasonal ramping of nest temperatures also caused shorter relative incubation periods in the later, all-female clutches. As a consequence of this developmental ‘catch-up’, the mean hatching date for male-producing nests preceded the mean hatching date for female-producing nests by only 7.2 days. We suggest that a contracted distribution of hatching dates compared to the distribution of oviposition dates represents a general pattern for oviparous reptiles in seasonal climates, which in TSD species may largely offset the temporal disparity in nesting dates between the sexes. Although data are needed for other TSD species, such minor age differences between male and female hatchlings may not translate into fitness differences later in life, an assumption of some models for the evolution and maintenance of TSD.     

No seasonal sex-ratio shift despite sex-specific fitness returns of hatching date in a lizard with genotypic sex determination

Sex allocation theory predicts that mothers should adjust their sex-specific reproductive investment in relation to the predicted fitness returns from sons versus daughters. Sex allocation theory has proved to be successful in some invertebrate taxa but data on vertebrates often fail to show the predicted shift in sex ratio or sex-specific resource investment. This is likely to be partly explained by simplistic assumptions of vertebrate life-history and mechanistic constraints, but also because the fundamental assumption of sex-specific fitness return on investment is rarely supported by empirical data. In short-lived species, the time of hatching or parturition can have a strong impact on the age and size at maturity. Thus, if selection favors adult sexual-size dimorphism, females can maximize their fitness by adjusting offspring sex over the reproductive season. We show that in mallee dragons, Ctenophorus fordi, date of hatching is positively related to female reproductive output but has little, if any, effect on male reproductive success, suggesting selection for a seasonal shift in offspring sex ratio. We used a combination of field and laboratory data collected over two years to test if female dragons adjust their sex allocation over the season to ensure an adaptive match between time of hatching and offspring sex. Contrary to our predictions, we found no effect of laying date on sex ratio, nor did we find any evidence for within-female between-clutch sex-ratio adjustment. Furthermore, there was no differential resource investment into male and female offspring within or between clutches and sex ratios did not correlate with female condition or any partner traits. Consequently, despite evidence for selection for a seasonal sex-ratio shift, female mallee dragons do not seem to exercise any control over sex determination. The results are discussed in relation to potential constraints on sex-ratio adjustment, alternative selection pressures, and the evolution of temperature-dependent sex determination.     

Do operational sex ratios influence sex allocation in viviparous lizards with temperature‐dependent sex determination?

Under certain environmental situations, selection may favour the ability of females to adjust the sex ratio of their offspring. Two recent studies have suggested that viviparous scincid lizards can modify the sex ratio of the offspring they produce in response to the operational sex ratio (OSR). Both of the species in question belong to genera that have also recently been shown to exhibit temperature-dependent sex determination (TSD). Here we test whether pregnant montane water skinks (Eulamprus tympanum) utilise TSD to select offspring sex in response to population wide imbalances in the OSR, by means of active thermoregulation. We use a combination of laboratory and field-based experiments, and conduct the first field-based test of this hypothesis by maintaining females in outdoor enclosures of varying OSR treatments throughout pregnancy. Although maternal body temperature during pregnancy was influenced by OSR, the variation in temperature was not great enough to affect litter sex ratios or any other phenotypic traits of the offspring.     

Does sex-ratio selection influence nest-site choice in a reptile with temperature-dependent sex determination?

Evolutionary theory predicts that dioecious species should produce a balanced primary sex ratio maintained by frequency-dependent selection. Organisms with environmental sex determination, however, are vulnerable to maladaptive sex ratios, because environmental conditions vary spatio-temporally. For reptiles with temperature-dependent sex determination, nest-site choice is a behavioural maternal effect that could respond to sex-ratio selection, as mothers could adjust offspring sex ratios by choosing nest sites that will have particular thermal properties. This theoretical prediction has generated decades of empirical research, yet convincing evidence that sex-ratio selection is influencing nesting behaviours remains absent. Here, we provide the first experimental evidence from nature that sex-ratio selection, rather than only viability selection, is probably an important component of nest-site choice in a reptile with temperature-dependent sex determination. We compare painted turtle (Chrysemys picta) neonates from maternally selected nest sites with those from randomly selected nest sites, observing no substantive difference in hatching success or survival, but finding a profound difference in offspring sex ratio in the direction expected based on historical records. Additionally, we leverage long-term data to reconstruct our sex ratio results had the experiment been repeated in multiple years. As predicted by theory, our results suggest that sex-ratio selection has shaped nesting behaviour in ways likely to enhance maternal fitness.     

Predicting the fate of a living fossil: how will global warming affect sex determination and hatching phenology in tuatara?

How will climate change affect species'' reproduction and subsequent survival? In many egg-laying reptiles, the sex of offspring is determined by the temperature experienced during a critical period of embryonic development (temperature-dependent sex determination, TSD). Increasing air temperatures are likely to skew offspring sex ratios in the absence of evolutionary or plastic adaptation, hence we urgently require means for predicting the future distributions of species with TSD. Here we develop a mechanistic model that demonstrates how climate, soil and topography interact with physiology and nesting behaviour to determine sex ratios of tuatara, cold-climate reptiles from New Zealand with an unusual developmental biology. Under extreme regional climate change, all-male clutches would hatch at 100% of current nest sites of the rarest species, Sphenodon guntheri, by the mid-2080s. We show that tuatara could behaviourally compensate for the male-biasing effects of warmer air temperatures by nesting later in the season or selecting shaded nest sites. Later nesting is, however, an unlikely response to global warming, as many oviparous species are nesting earlier as the climate warms. Our approach allows the assessment of the thermal suitability of current reserves and future translocation sites for tuatara, and can be readily modified to predict climatic impacts on any species with TSD.     

Exploring the evolution of environmental sex determination, especially in reptiles

Environmental sex determination has been documented in a variety of organisms for many decades and the adaptive significance of this unusual sex-determining mechanism has been clarified empirically in most cases. In contrast, temperature-dependent sex determination (TSD) in amniote vertebrates, first noted 40 years ago in a lizard, has defied a general satisfactory evolutionary explanation despite considerable research effort. After briefly reviewing relevant theory and prior empirical work, we draw attention to recent comparative analyses that illuminate the evolutionary history of TSD in amniote vertebrates and point to clear avenues for future research on this challenging topic. To that end, we then highlight the latest empirical findings in lizards and turtles, as well as promising experimental results from a model organism, that portend an exciting future of progress in finally elucidating the evolutionary cause(s) and significance of TSD.     

Communal Nesting in Asocial Abert's Squirrels: the Role of Social Thermoregulation and Breeding Strategy

The social thermoregulation hypothesis states that endothermic species will communally nest to reduce energy expenditures on thermoregulation. The hypothesis predicts that the frequency of communal nesting should increase with decreasing ambient temperature. The potential costs of communal nesting (e.g., increased predation risk, resource competition, cuckoldry, parasite/disease transmission, or infanticide) could decrease the frequency of communal nesting especially for asocial breeding females with dependent offspring. We examined the effects of ambient temperature and seasonal reproductive activities on the probability of communal nesting in Abert's squirrels (Sciurus aberti) in the Pinaleño Mountains, Arizona. Most squirrels nested consistently with the same partner in mixed‐sex pairs. The proportion of individuals engaging in communal nesting increased with decreasing ambient temperature as predicted by the social thermoregulation hypothesis. The onset of the breeding season greatly reduced the proportion of individuals communally nesting. The negative relationship between ambient temperature and communal nesting supports the use of communal nesting in Abert's squirrels as a mechanism to reduce thermoregulatory costs during cold conditions. The abrupt drop in the frequency of communal nesting during the breeding season is likely due to female abandonment of this behavior. By avoiding communally nesting during the breeding season, females may prevent males from mating with them outside of mating chases, reduce resource competition, and protect offspring from infanticide, diseases, and parasites. Males may gain additional fitness benefits from nesting with females because familiarity with females increases dominance rank in mating activities.     

Nest site choice compensates for climate effects on sex ratios in a lizard with environmental sex determination

Theoretical models suggest that in changing environments natural selection on two traits, maternal nesting behaviour and pivotal temperatures (those that divide the sexes) is important for maintaining viable offspring sex ratios in species with environmental sex determination (ESD). Empirical evidence, however, is lacking. In this paper, we provide such evidence from a study of clinal variation in four sex-determining traits (maternal nesting behaviour, pivotal temperatures, nesting phenology, and nest depth) in Physignathus lesueurii, a wide-ranging ESD lizard inhabiting eastern Australia. Despite marked differences in air and soil temperatures across our five study sites spanning 19° latitude and 1200 m in elevation, nest temperatures did not differ significantly among sites. Lizards compensated for climatic differences chiefly by selecting more open nest sites with higher incident radiation at cooler sites. Clinal variation in the onset of nesting also compensated for climatic differences, but to a lesser extent. There was no evidence of compensation through pivotal temperatures or nest depth. More broadly, our results extend to the egg stage the life history prediction that behaviour is the chief compensatory mechanism for climatic differences experienced by species spanning environmental extremes. Furthermore, our study was unique in revealing that nest site choice influenced mainly the daily range in nest temperatures, rather than mean temperatures, in a shallow-nesting reptile. Finally, indirect evidence suggests that the cue used by nesting lizards was radiation or temperature (through basking or assessing substrate temperatures), not visual detection of canopy openness. We conclude that maternal nesting behaviour and nesting phenology are traits subject to sex ratio selection in P. lesueurii, and thus, must be considered among the repertoire of ESD species for responding to climate change.     

Genetic evidence for co-occurrence of chromosomal and thermal sex-determining systems in a lizard

An individual's sex depends upon its genes (genotypic sex determination or GSD) in birds and mammals, but reptiles are more complex: some species have GSD whereas in others, nest temperatures determine offspring sex (temperature-dependent sex determination). Previous studies suggested that montane scincid lizards (Bassiana duperreyi, Scincidae) possess both of these systems simultaneously: offspring sex is determined by heteromorphic sex chromosomes (XX-XY system) in most natural nests, but sex ratio shifts suggest that temperatures override chromosomal sex in cool nests to generate phenotypically male offspring even from XX eggs. We now provide direct evidence that incubation temperatures can sex-reverse genotypically female offspring, using a DNA sex marker. Application of exogenous hormone to eggs also can sex-reverse offspring (oestradiol application produces XY as well as XX females). In conjunction with recent work on a distantly related lizard taxon, our study challenges the notion of a fundamental dichotomy between genetic and thermally determined sex determination, and hence the validity of current classification schemes for sex-determining systems in reptiles.     

Segregating variation for temperature-dependent sex determination in a lizard

Temperature-dependent sex determination (TSD) was first reported in 1966 in an African lizard. It has since been shown that TSD occurs in some fish, several lizards, tuataras, numerous turtles and all crocodilians. Extreme temperatures can also cause sex reversal in several amphibians and lizards with genotypic sex determination. Research in TSD species indicates that estrogen signaling is important for ovary development and that orthologs of mammalian genes have a function in gonad differentiation. Nevertheless, the mechanism that actually transduces temperature into a biological signal for ovary versus testis development is not known in any species. Classical genetics could be used to identify the loci underlying TSD, but only if there is segregating variation for TSD. Here, we use the ‘animal model'' to analyze inheritance of sexual phenotype in a 13-generation pedigree of captive leopard geckos, Eublepharis macularius, a TSD reptile. We directly show genetic variance and genotype-by-temperature interactions for sex determination. Additive genetic variation was significant at a temperature that produces a female-biased sex ratio (30 °C), but not at a temperature that produces a male-biased sex ratio (32.5 °C). Conversely, dominance variance was significant at the male-biased temperature (32.5 °C), but not at the female-biased temperature (30 °C). Non-genetic maternal effects on sex determination were negligible in comparison with additive genetic variance, dominance variance and the primary effect of temperature. These data show for the first time that there is segregating variation for TSD in a reptile and consequently that a quantitative trait locus analysis would be practicable for identifying the genes underlying TSD.     

Temperature-Dependent Sex Determination in Sea Turtles in the Context of Climate Change: Uncovering the Adaptive Significance

The adaptive significance of temperature-dependent sex determination (TSD) in reptiles remains unknown decades after TSD was first identified in this group. Concurrently, there is growing concern about the effect that rising temperatures may have on species with TSD, potentially producing extremely biased sex ratios or offspring of only one sex. The current state-of the-art in TSD research on sea turtles is reviewed here and, against current paradigm, it is proposed that TSD provides an advantage under warming climates. By means of coadaptation between early survival and sex ratios, sea turtles are able to maintain populations. When offspring survival declines at high temperatures, the sex that increases future fecundity (females) is produced, increasing resilience to climate warming. TSD could have helped reptiles to survive mass extinctions in the past via this model. Flaws in research on sex determination in sea turtles are also identified and it is suggested that the development of new techniques will revolutionize the field.     

The role of Haldane's rule in sex allocation

Sex allocation theory predicts that parents should bias their reproductive investments toward the offspring sex generating the greatest fitness return. When females are the heterogametic sex (e.g., ZW in butterflies, some lizards, and birds), production of daughters is associated with an increased risk of offspring inviability due to the expression of paternal, detrimental recessives on the Z chromosome. Thus, daughters should primarily be produced when mating with partners of high genetic quality. When female sand lizards (Lacerta agilis) mate with genetically superior males, exhibiting high MHC Class I polymorphism, offspring sex ratios are biased towards daughters, possibly due to recruitment of more Z-carrying oocytes when females have assessed the genetic quality of their partners. If our study has general applicability across taxa, it predicts taxon-specific sex allocation effects depending on which sex is the heterogametic one.     

Open-cell parasitism shapes maternal investment patterns in the Red Mason bee Osmia rufa

Brood cell parasitism inflicts high fitness costs on solitary,nest-constructing bees. Many of these parasites enter open cellsduring its provisioning, when the mother bee is absent. Therefore,females can reduce the risk of open-cell parasitism by limitingthe time they are away from the nest. However, provisioningefficiency (provisioning time per unit of progeny body mass)decreases due to aging. To limit the increasing risk of open-cellparasitism as the nesting season progresses, female bees couldoptimize their maternal investment strategy by shifting thesex ratio and the body size of offspring during the nestingseason. This prediction was tested in the Red Mason bee Osmiarufa (O. bicornis), a stem- or hole-nesting, polylectic, univoltinemegachilid bee. In O. rufa, the risk of open-cell parasitismwas found to be correlated with cell provisioning time. Additionally,the provisioning efficiency of females declined during the nestingseason to one-fourth of the initial value. However, cell-provisioningtime did not increase correspondingly. Bees dealt with theirdecreasing provisioning efficiency by reducing the amount ofstored larval food, leading to a reduction of offspring sizeand a seasonal shift toward males in the offspring sex ratio.The influence of provisioning efficiency and risk of open-cellparasitism on optimal offspring size was analyzed by means ofa statistical model. The observed maternal investment patternof Red Mason bees is an adaptive strategy to reduce open-cellparasitism.     

Change in offspring sex ratio over a very short season in Lincoln's Sparrows: the potential role of bill development

ABSTRACT The sex ratios of offspring are targets of natural selection that can affect parental energy expenditure and fitness, adult sex ratios, and population dynamics. Parents may manipulate offspring sex ratios based on sex differences in their offsprings' potential for reproductive success. In Lincoln's Sparrows (Melospiza lincolnii), male bill shape is associated with the quality of songs, and song quality predicts female preferences in a reproductive context. Males and females that hatch later relative to brood mates or later in the breeding season tend to develop bill shapes that are, for males, associated with low‐quality song. Because females do not sing and do not experience this selection pressure, we predicted that the sex of offspring produced late relative to their brood mates or relative to the season should be biased toward females. Using a molecular technique to sex nestlings, we found no effects of hatching order or any interaction between date of clutch initiation (season) and hatching order on offspring sex. However, we found a seasonal decline in the proportion of male offspring, from approximately 0.8 at the beginning to 0.4 at the end of a clutch initiation season only 19 d in duration. To our knowledge, this is the shortest period over which the offspring sex ratio has been shown to change in a bird population. Moreover, these findings are consistent with the hypothesis that sex differences in the potential attractiveness of offspring ultimately influence offspring sex ratios.     

Unexpected resilience of species with temperature-dependent sex determination at the Cretaceous-Palaeogene boundary

It has been suggested that climate change at the Cretaceous-Palaeogene (K-Pg) boundary, initiated by a bolide impact or volcanic eruptions, caused species with temperature-dependent sex determination (TSD), including dinosaurs, to go extinct because of a skewed sex ratio towards all males. To test this hypothesis, the sex-determining mechanisms (SDMs) of Cretaceous tetrapods of the Hell Creek Formation (Montana, USA) were inferred using parsimony optimizations of SDMs on a tree, including Hell Creek species and their extant relatives. Although the SDMs of non-avian dinosaurs could not be inferred, we were able to determine the SDMs of 62 species; 46 had genotypic sex determination (GSD) and 16 had TSD. The TSD hypothesis for extinctions performed poorly, predicting between 32 and 34 per cent of survivals and extinctions. Most surprisingly, of the 16 species with TSD, 14 of them survived into the Early Palaeocene. In contrast, 61 per cent of species with GSD went extinct. Possible explanations include minimal climate change at the K-Pg, or if climate change did occur, TSD species that survived had egg-laying behaviour that prevented the skewing of sex ratios, or had a sex ratio skewed towards female rather than male preponderance. Application of molecular clocks may allow the SDMs of non-avian dinosaurs to be inferred, which would be an important test of the pattern discovered here.