Effects of an immune challenge on the thermal preferences of adult and newborn Liolaemus lizards from Patagonia, Argentina.

Body temperature has relevant effects on the immune response. Here, we characterized the thermal biology and health condition of the viviparous lizard Liolaemus kingii from Patagonia (Argentina), by studying field body temperatures, presence of injuries or ectoparasites, body condition (BC), and individual immune response capacity with the phytohemagglutinin (PHA) skin-swelling assay. In addition, we analyzed the effects of injections of a bacterial endotoxin (lipopolysaccharide; LPS) on the preferred temperature (Tp) and BC of adult males and newborns. The PHA treatment caused detectable thickening at 2 and 20 hours post-assay in males, indicating a significant immune response related to an increase in cellular activity. LPS-challenged lizards thermoregulated accurately and at stable body temperatures within the 50% interquartile of Tp (Tset) over the 72-hour period while the control group showed a more variable and lower Tp. Exposure to LPS negatively affected the BC of newborns, whereas it did not affect the BC of adult males. LPS challenges, used as a proxy of pathogen exposures to study lizard behavioral thermoregulation, constitute a practical approach to assess the immunological constraints lizards from high-latitude regions may face due to global warming and anthropogenic disturbances.

Thermophysiological plasticity could buffer the effects of global warming on a Patagonian lizard.

Ecophysiological plasticity determines, to a great extent, the geographic distribution and the vulnerability of ectotherms to climate change. We studied the relationship between locomotor performance and temperature of Liolaemus elongatus lizards in three populations in northern Patagonia, Argentina, differing in thermal characteristics. We related the thermophysiological and locomotor performance parameters with the environmental conditions currently experienced by these populations and analyzed whether the expected increment of the environmental temperature due to climate change could affect these vital traits. We also determined, for one of the populations, the effects of 30 acclimation days at two temperature treatments (22°C and 30°C) on running speed, thermal preference in the laboratory (Tpref ), panting threshold, and minimum critical temperature. We found that L. elongatus, despite the differences in environmental temperatures among the three sites, exhibited maximum speed at similar temperatures (optimum temperature for locomotor performance; To ). The southern populations currently experience temperatures below that required to reach their maximum locomotor performance while the northernmost population is threatened by peaks of high temperatures that exceed the To . Therefore, global warming could diminish lizards' running performance in northern populations and lizards may spend more time refuging and less time on other activities such as feeding, territory defense, and dispersion. However, we show evidence of plasticity in L. elongatus locomotor performance when acclimated at high temperatures resulting in a potential advantage to cushion the effect of the rising environmental temperatures expected during climate change.

Wintertime tales: How the lizard Liolaemus lineomaculatus endures the temperate cold climate of Patagonia, Argentina.

In temperate, polar and montane environments, ectotherms must find ways to endure throughout the coldest months of the year. Lizards search for microsites where temperatures remain warm or alter their biochemical balance to tolerate freezing or avoid it by supercooling. We evaluated the cold hardiness and potential winter refuges of two populations of Liolaemus lineomaculatus, from a temperate site (42°S) and a cold site (50°S). We analysed the role of possible cryoprotectants by comparing a group of cooled-down lizards with a control group of lizards that were not exposed to cold. The populations of this study are not freeze tolerant and the biochemical analysis showed no evidence of metabolites significantly changing concentration after exposure to cold. However, the species remained several hours at their Supercooling Point (SCP), suggesting they can supercool. The analysis of potential winter refuges showed that lizards using these potential refuges would spend almost no time at all at temperatures close to or below their SCP. Furthermore, lizards from the cold site were able to survive below 0°C temperatures with a lower SCP than lizards from the temperate site. Liolaemus lineomaculatus developed physiological mechanisms that can help them survive when temperatures drop sharply, even when lizards are in suitable shelters.

Thermal biology and locomotor performance in Phymaturus calcogaster: are Patagonian lizards vulnerable to climate change?

Behavioral and physiological traits of ectotherms are especially sensitive to fluctuations of environmental temperature. In particular, niche-specialist lizards are dependent on their physiological plasticity to adjust to changing environmental conditions. Lizards of the genus Phymaturus are viviparous, mainly herbivorous, and inhabit only rock promontories in the steppe environments of Patagonia and the Andes. Herein, we examine the vulnerability of the southernmost Phymaturus species to global warming: the endemic Phymaturus calcogaster, which lives in a mesic environment in eastern Patagonia. We studied body temperatures in the field (Tb ), preferred body temperatures in a thermogradient (Tpref ), the operative (Te ) and environmental temperatures, and the dependence of running performance on body temperature. P. calcogaster had a mean Tb (27.04°C) and a mean Te (31.15°C) both lower than their preferred temperature (Tpref = 36.61°C) and the optimal temperature for running performance (To = 37.13°C). Lizard activity seems to be restraint during the early afternoon due high environmental temperatures. However, both, the high safety margin and warming tolerance suggest that the expected increase in environmental temperatures due to global warming (IPCC report in 2018) would not threaten, but indeed enhance locomotor performance in this population.

Decrease in preferred temperature in response to an immune challenge in lizards from cold environments in Patagonia, Argentina.

In ectotherms, the likelihood of surviving an infection is determined by the efficiency of thermoregulation, the availability of a variety of thermal microenvironments, the individual's health status, and the virulence of the infective agent. Physiological and behavioral demands related to an efficient immune response entail a series of costs that compete with other vital activities, specifically energy storage, growth, reproduction, and maintenance functions. Here, we characterize the thermal biology and health status by the presence of injuries, ectoparasites, body condition, and individual immune response capacity (using phytohemagglutinin in a skin-swelling assay) of the southernmost lizards of the world, Liolaemus sarmientoi, endemic to a sub-optimal, cold environment in Patagonia, Argentina. In particular, we study the effect of a bacterial endotoxin (lipopolysaccharide; LPS-treatment) on thermoregulation. We found that the field-active body temperature (Tb) was much lower than the preferred body temperature (Tp) obtained in the laboratory. All the individuals were in good body condition at the beginning of the experiments. The phytohemagglutinin test caused detectable thickening in sole-pads at 2 h and 24 h post-assay in males and non-pregnant females, indicating a significant innate immune response. In the experimental immune challenge, the individuals tended to prefer a low body temperature after LPS-treatment (2 h post-injection) and developed hypothermia, while the control individuals injected with phosphate buffered saline (PBS), maintained their body temperature throughout the trial. In both the LPS-treatment and PBS-control individuals, BC declined during the experiment. Hypothermia may allow this southernmost species to optimize the use of their energetic resources and reduce the costs of thermoregulation in a cold-temperate environment where they rarely attain the mean Tp (35.16 °C) obtained in laboratory.

Impact of temperature on bite force and bite endurance in the leopard iguana (Diplolaemus leopardinus) in the Andes Mountains.

In ectotherms, temperature exerts a strong influence on the performance of physiological and ecological traits. One approach to understanding the impact of rising temperatures on animals and their ability to cope with climate change is to quantify variation in thermal-sensitive traits. Here, we examined the thermal biology, temperature dependence and thermal plasticity of bite force (endurance and magnitude) in Diplolaemus leopardinus, an aggressive and territorial lizard endemic to Mendoza province, Argentina. Our results indicate that this lizard behaves like a moderate thermoregulator that uses the rocks of its environment as the main heat source. Bite endurance was not influenced by head morphometry and body temperature, whereas bite force was influenced by head length and jaw length, and exhibited thermal dependence. Before thermal acclimation treatments, the maximum bite force for D. leopardinus occurred at the lowest body temperature and fell sharply with increasing body temperature. After acclimation treatments, lizards acclimated at higher temperatures exhibited greater bite force. Bite force showed phenotypic plasticity, which reveals that leopard iguanas are able to maintain (and even improve) their bite force under a rising-temperature scenario.

Impact of immunological state on eco-physiological variables in one of the southernmost lizards in the world.

The immune state is an essential component of survival as it directly influences physiological performance and health status. Variation in the leukocyte profile, a significantly increase in body temperature, and a detriment of the eco-physiological performance are among the possible consequences of an unhealthy state. In this study we analyse and discuss how field body temperature, preferred body temperature, the speed for sprint and long runs, locomotor stamina, and body condition can be affected by the immunological state (i.e. leukocyte profile) in a wild population of Liolaemus sarmentoi. Juveniles and adult males with a high percentage of eosinophils, basophils, and a low percentage of monocytes preferred higher body temperatures in a thermal gradient, while pregnant females maintained thermal preferences independently of leukocyte profile. Although juveniles with a high percentage of heterophils showed less locomotor stamina, adult males and pregnant females showed no differences in locomotor performance in relation to leukocyte profile. This study represents a starting point in eco-immunology of a wild lizard population of Liolaemus in cold and temperate environments of Patagonia where the southward shift in the geographic ranges of pathogen populations due to global warming represents a threat to resident host populations.

Reproductive biology of a viviparous lizard (Mabuya dorsivittata) from the subtropical Wet Chaco of Argentina: geographical variations in response to local environmental pressures.

Herein we studied the reproductive biology of a viviparous lizard (Mabuya dorsivittata) from the Wet Chaco region (northeastern Argentina) and compared the results with other populations from the Espinal (central Argentina) and the Atlantic Forest (southeastern Brazil), and with other Neotropical species of Mabuya to better understand the possible causes of its reproductive phenotype variation. Males and females of M. dorsivittata from the Wet Chaco exhibited associated, seasonal, and annual reproductive cycles. Spermatogenic activity related positively to a lengthening photoperiod reaching maximum activity in late spring (December). Females displayed an extended gestation period of 11 months, from mid-summer (February) to late spring or early summer (December) when births occur. Embryonic development was associated with temperature and historical rainfall. Litter size ranged from 3 to 8 (mean = 5.3 ± 1.3 SD) and increased with body size and body mass of females. Fat-body mass varied seasonally and was inversely correlated with spermatogenesis and to embryonic development. Females were larger in body size and interlimb length, and smaller in head length than males. We observed interpopulational differences in minimum body size, litter size, and timing of birth, probably as a result of phenotypic plasticity, genetic divergence or both.

Effects of Climate and Latitude on Age at Maturity and Longevity of Lizards Studied by Skeletochronology.

Longevity and age at maturity are key life-history traits, directly linked to fitness attributes such as survival and reproductive output. It has been proposed that these traits are strongly influenced by environmental factors, such as temperature, seasonality, and precipitations, which determine the existence of a continuum of life-histories that goes from the "slow" life histories characterized by late maturity and high longevity of cold and highly seasonal climates to the "fast" life histories characterized by early maturity and low longevity, typical of the tropical climates. However, large-scale studies that address these topics in lizards are scarce and most of them are based on heterogeneous data, which may overlook the real patterns. Using skeletochronology, we studied age at maturity and longevity of two species of Phymaturus lizards, Phymaturus aguanegra from the Andes and Phymaturus zapalensis from the Patagonian steppe (Argentina). Then, we confronted longevity and age at maturity in these species with published skeletochronology-based data on 46 other lizard species to examine possible association of these life-history traits with latitude and mean annual temperature, thermal amplitude, and precipitations. Both Phymaturus species showed late sexual maturity (7 and 8-9 years, respectively) and high longevity (16 and 14-15 years, respectively) in coincidence with the other species of the genus studied up to date. The phylogenetic comparative analysis revealed that the most important variable in the determination of longevity patterns in the species studied was latitude: at higher latitudes lizards tend to live longer. In contrast, age at sexual maturity was dependent on mean annual temperature most, especially in males, as lizards from hotter climates mature earlier than lizards from cold sites.

Behavioral and physiological polymorphism in males of the austral lizard Liolaemus sarmientoi.

Integrative behavioral studies show that the interplay between individual physiology and social behavior influences the ecology of the species, ultimately affecting individual fitness. Particularly in lizards, color polymorphism is associated with differential behaviors and reproductive strategies, which are evident in mature males during the mating season. Dominant males generally have greater endurance, higher body temperature, and larger bodies than submissive males, so they can acquire and defend larger territories and have greater access to females for mating. We studied whether the color morphs observed in males of one of the world's southernmost reptiles, Liolaemus sarmientoi, are related to behavioral variation during agonistic interactions, thermal physiology, morphology, and/or locomotor stamina. Liolaemus sarmientoi males exhibit three color morphs: red (RR), red-yellow (RY), and yellow (YY). These lizards exhibit subtle behavioral displays and we did not observe stamina differences among morphs. However, we found that RR males are more aggressive than YY males during agonistic encounters. In addition, greater body temperature change during trials, higher field body temperatures, and greater head sizes of RR males compared to RY or YY indicate that RR is a dominant morph, which may influence their ability to acquire and defend territory and tactics for achieving reproductive success.

Slow life histories in lizards living in the highlands of the Andes Mountains.

In the highlands of the Andes, lizards must balance precisely the allocation of energy for growth and reproduction to ensure their survival. We studied the individuals' age, growth rates, age at sexual maturity, and maximum life span of the viviparous lizard Phymaturus antofagastensis, endemic of cold and harsh environments at high altitudes in the Andes Mountains of Catamarca province, Argentina. We also estimated key life history parameters like reproductive effort, lifetime reproductive effort, net reproductive rate, and relative reproductive time in P. antofagastensis as well as in other Phymaturus to compare the interplay among growth, maintenance, and reproduction in species that live across a latitudinal and altitudinal gradient. We found that females and males of P. antofagastensis mature late in life, at 6-7 years old, respectively, and some individuals reached 20 years of age. Adult females showed higher specific growth rates than males and an adult life span of 9 years which, due to their biennial reproduction, results in an estimated production of only four litters in life. This species exhibits one of the highest lifetime reproductive efforts described for lizards. Our results indicate the existence of a tradeoff between the number of reproductive events throughout life and reproductive effort devoted to each event in Phymaturus, related to the phylogenetic group. The palluma group shows low reproductive effort but high number of reproductive events throughout their lives, whereas the patagonicus group shows high reproductive efforts in low number of reproductive events.

How does a viviparous semifossorial lizard reproduce? Ophiodes intermedius (Squamata: Anguidae) from subtropical climate in the Wet Chaco region of Argentina.

The best predictors of reproductive patterns are commonly associated with climate factors, but evolutionary history also plays an important role. The semifossorial and viviparous lizard Ophiodes intermedius from the Wet Chaco region of Argentina showed an annual cycle with asynchrony between males and females and an unusual pattern for subtropical climates, with vitellogenesis beginning in autumn, ovulation and copulation in spring, and births occurring in summer. Males exhibited annual variation of testicular size associated with spermatogenic activity, reaching their maximum gonadal activity in late summer (March), but sperm storage in the epididymis and/or deferent duct occurred throughout the year. Females showed an extended reproductive cycle beginning in mid-autumn (May) with vitellogenesis and finishing with births from late spring to mid-summer (December to February). Litter size varied from 4 to 9 offspring. Females reached sexual maturity at a larger snout-vent length and, overall, showed greater body size than males, while males exhibited larger heads than females. Fat body cycles indicated that females use lipid reserves to support vitellogenesis and embryo development, while males allocate lipid resources to the search for females, courtship and copulation rather than to gametogenesis. Ophiodes intermedius differed from other species of the genus in litter size, gestation period, timing of birth and the minimum size at sexual maturity, probably as a result of the influence of ecological, historical and phylogenetic factors.

Reproductive strategies in males of the world's southernmost lizards.

Reproductive and life history patterns in reptiles are tightly related to the environmental conditions, so male reproductive cycles have been historically characterized as continuous, for tropical lizards, or seasonal, for temperate lizards. However, males of Liolaemus and Phymaturus lizards (Liolaemidae), from cold temperate climates of high altitudes or latitudes in Argentina and Chile, have developed a variety of reproductive cycles to coordinate with the short female reproductive season and to deal with the low frequency of reproductive females in the population. Using gonadal histology and morphological analysis, we describe the male reproductive biology, fat storage and sexual dimorphism of the viviparous lizards Liolaemus sarmientoi and Liolaemus magellanicus that inhabit an austral grass steppe at 51°S, in the southern limit of the American continent. Males of L. sarmientoi and L. magellanicus are reproductively available during the entire activity season of approximately 5 months. In addition, males of both species exhibit greater body sizes than females in morphological variables relevant in sexual selection. Meanwhile, females of both species exhibit larger inter-limb length than conspecific males, which suggests fecundity selection to increase space for a larger litter size. The continuous sperm production throughout the activity season allows these liolaemids to mate at any time when females ovulate, representing a selective advantage to deal with the short activity season and the adversities of the cold environment they inhabit.

Contributions to the study of oviparity-viviparity transition: placentary structures of Liolaemus elongatus (Squamata: Liolaemidae).

Liolaemus elongatus (Liolaemidae) is a viviparous, mainly lecithotrophic species with placental structures specialized for uptake of oxygen and inorganic nutrient transport. An allantoplacenta and an omphaloplacenta are present during early embryonic stages (25-28) and there is a moderate distension of the uterine wall and major glandular activity in the uterine mucosa and submucosa compared with nonpregnant females. The uterine epithelium increases in height, first as a growth in the height of some dispersed cells localized in all regions of the placenta, and later as groups of cells localized in the periembryonic and central-abembryonic regions. At embryonic stage 39, the allantoplacenta reaches its maximum extension around the yolk sac. Omphaloplacenta is restricted to the abembryonic zone, the yolk cleft limiting the newly formed isolated vitelline mass. At more advanced embryonic stages (39-42), the blood supply to the allantoplacenta's periembryonic zone increases, matching the profuse allantoic vascularization. At embryonic stage 42, a secondary cleft opens in the main vitelline mass, above the first yolk cleft, and allantoic blood vessels enter into this secondary cleft. This secondary cleft subdivides the vitelline mass into a large embryonic region connected to a much smaller abembryonic region. In L. elongatus most nutritional resources seem to be provided by the yolk that remains attached to the newborn for 2 or 3 days as an external supply. The embryo's wet weight doubles the weight of the decrease observed in vitelline mass. But the dry wet diminishes, evidencing the importance of the exchange of water and inorganic nutrients.