Selection on body size tends to favor larger males that outcompete smaller males to mate with females, and larger, more fecund females. For many web-building spiders in the Nephilidae family, reproductive success increases with body size, which in turn, is related to diet. The diet of female spiders may overlap with males who share her web, but diet patterns could depend on size if certain males have better access to prey ensnared in the web. Therefore, we tested the hypothesis that Joro spider (Trichonephila clavata) dietary patterns are sex-specific and related to body size and condition. We combined morphometric size analysis with molecular gut content analysis via DNA metabarcoding and high-throughput sequencing to characterize male and female Joro spider diets from 60 webs at 25 locations. Female Joro spiders were larger than males and their diets were more diverse. While male Joro spiders consumed the same prey taxa as females, more than 50 additional taxa were detected in female diets, which led to significant dissimilarity in prey composition between sexes. As male body size increased, diet diversity and body condition decreased, suggesting a potential trade-off between time spent foraging versus active defense of their position on the web. Female and male spiders captured from the same web shared prey taxa. However, the proportion of prey overlap was not related to male size. Combined, Joro spiders show sex-specific dietary patterns, and males appear to have less access to the diverse food captured in female webs.

The study of animals’ activity and behavior in the wild is an extremely challenging task. Although tri-axial accelerometers are invaluable for behavioral analyses, their use is more frequent in large charismatic endotherms with limited application in ectotherms. The scarce utilization of this methodology on small-size reptiles is focused on animals’ activity and energetics, showing few records of rapid displays and behavior signals. Here, we present a novel multidisciplinary approach capable of advancing research on reptiles’ behavior. Our proposed approach uses advanced technologies for the digitization, reconstruction and visualization of reptiles and their behavior. We (i) record movement through tri-axial accelerometers, video cameras, and motion capture systems; (ii) ground-truth data through the video records; (iii) develop realistically accurate 3D avatars of the recorded movement for visualization purposes, and (iv) archive data on a Behavior Pattern Database. As case studies, we used two small Mediterranean reptiles, the lizard Laudakia cypriaca and the snake Dolichophis jugularis. Through our approach, we successfully recorded, ground-truthed, and labeled for the first time, several detailed movements and behaviors of the two case study species. We developed an accurate digital overview of those movements using motion capture and 3D animal reconstruction. Finally, we structured a database for archiving all behavioral data and demonstrated how those archives can be used for advancing behavioral research, providing ecological insights into this animal group. Our approach can enhance research on reptiles’ behavior by contributing to the analysis of complex or isolated behaviors, poorly studied, such as signals and social interactions, providing valuable insights and assisting behavioral analysis.

Sperm transfer constraints are common phenomena in spider mating systems, influenced by both behavioral and morphological factors. Non-motile, encapsulated sperm cells must navigate through narrow male and female ducts during short (only seconds) copulations, all while under the pressure of female aggression, which can include sexual cannibalism and genital damage. However, little attention has been paid to the impact of male and female internal genital morphology on sperm transfer. In this study, we quantify the number of sperm transferred by males in five species of the genus Argiope as a direct measure of reproductive success. We explore sperm transfer in relation to copulation duration, sperm availability, sexual dimorphism, and female aggression. Additionally, we compare intra- and interspecific variation in sperm transfer by examining internal male and female genital morphological traits. Finally, we discuss the allometry of internal genital morphology in terms of sperm transfer in spiders. Our findings indicate that the occurrence of sexual cannibalism is the significant factor impacting sperm transfer in the genus Argiope. We observed a positive, albeit non-significant, correlation between sperm duct width and sperm transfer. Nonetheless, interspecific variability in allometry and methodological challenges underscore the need for continued research to fully understand the complexities of internal genital evolution in spiders.

The burrow microhabitats created by burrowing mammals, as a hotspot for biodiversity distribution in ecosystems, provide multiple critical resources for many other sympatric species. However, the cascading effects of burrow resources on sympatric animal community assemblages and interspecific interactions are largely unknown. During 2020–2023, we monitored 184 Chinese pangolin (Manis pentadactyla) burrows using camera traps to reveal the burrow utilization patterns of commensal species. We totally recorded up to 57 species, with 19 mammal species, 32 bird species, and 1 reptile species recorded in the burrows revisited by Chinese pangolin, with 19 mammal species and 25 bird species in the non-revisited burrows. Among them, most bird species as peripheral species primarily utilize soil mounds while most mammal species as burrow-used species utilize burrow tunnels. The structure of animal communities in the burrows revisited by Chinese pangolins is more complex than that in the burrows not revisited. Furthermore, the positive correlation between community species in pangolin-revisited burrows is also stronger. Our results demonstrate that the presence and repeated visitation by Chinese pangolins could enhance positive interactions (i.e., the emergence of one species promotes the emergence of another) among species that utilize the burrow resources (particularly, burrow-used species). Our study provides the first evidence that the ecological role of the Chinese pangolin and its associated burrow microhabitats in promoting the coexistence of burrowing commensals and the restoration of Chinese pangolin populations may potentially contribute to the restoration of local biodiversity and ecological processes.

Primates demonstrate variability regarding the degree to which they display status hierarchies, which are influenced by a multitude of demographic and ecological factors. Additionally, primates must manage their interactions within these social hierarchies through the use of diverse communicative signals, including facial signals and manual gestures. Often times, these variables are assessed independently; however, it is probable that they collectively influence social rankings among primates. Our study investigates the application and accuracy of data-driven techniques, based on the genetic algorithm approach, in capturing social rankings among a group of captive chimpanzees, focusing on the analysis of communicative and demographic factors. We utilize observational data collected from a group of 18 chimpanzees residing at the Los Angeles Zoo from 2017 to 2019, derived from three previous studies carried out by the first author (BF). Our data-driven model exhibited a high degree of accuracy in capturing established social hierarchies in 2017, in addition to identifying notable fluctuations in rankings during periods of social instability from 2018 to 2019, especially in the aftermath of the passing of the highest-ranking female in the troop. Feature importance analysis revealed that social bond strength, measured via the dyadic composite sociality index (DCSI), was the most significant predictor of rank, highlighting the importance of social bonding in shaping status hierarchies. These models provide valuable insights for future research on primate behavior and social structures, as well as assist in making informed decisions for zoo management.

Animal signals are complex, comprising multiple components influenced by ecological factors and viewing perspectives that together impact their overall effectiveness. Our study explores how these factors affect the efficacy of multi-component signals in the Qinghai toad-headed agama, Phrynocephalus vlangalii. Using 3D animations, we simulated natural environments to evaluate how tail coiling and tail lashing—two primary tail displays—vary in effectiveness from both conspecific and predator perspectives under different ecological conditions. Baseline comparisons showed no significant difference in effectiveness between tail coiling and tail lashing without environmental constraints, though side-on tail coiling was consistently more effective than front-on displays. When noise proximity was introduced, tail lashing was more effective when the noise source was nearby, but this advantage diminished with distance. Conversely, tail coiling maintained consistent effectiveness across varying noise proximities, especially from a side-on view. In complex habitats with diverse plant species and varying wind conditions, tail lashing proved more effective, particularly from a front-on perspective, while tail coiling excelled from a side-on view. From a predator's perspective, tail lashing was slightly more effective under low wind conditions at close distances, though its visibility decreased with higher wind speeds. These findings highlight the adaptive significance of multi-component signals and the critical role of signal orientation in enhancing communication. This research offers insights into the evolutionary pressures shaping animal communication strategies.

On their dorsal surface, bat tongues show a raised muscular structure called mediodorsal lobe (MDL) or lingual prominence. It exhibits different mechanical papillae across species, which we hypothesized are linked to foraging strategies. We predicted that tall MDLs and prominent papillae pointing frontward would effectively trap prey items caught on the wing by aerial hawkers. We examined 904 high-resolution, close-up images of 239 individual bats from 24 European species, focusing on MDL height and morphology and arrangement of MDL papillae. Aerial hawkers such as Tadarida teniotis, Nyctalus lasiopterus, Miniopterus schreibersii, and pipistrelles displayed prominent forward-pointing papillae and taller MDLs, adaptations suited to high-speed aerial foraging. These traits may be part of a broader “ecomorphological syndrome”, facilitating efficient prey capture in open-space foragers. In contrast, gleaning and trawling species lacked these specializations, exhibiting flatter MDLs and less prominent papillae. Phylogenetic analysis indicated evolutionary convergence in MDL morphology among aerial hawkers, with M. schreibersii showing similarities to vespertilionids despite its phylogenetic distance. This convergence highlights the influence of evolutionary pressures arising from foraging requirements on tongue morphology. Although possessing tall MDLs, rhinolophids are outliers, possibly reflecting their unique perch-hunting strategy or echolocation. If the patterns we found are confirmed for a larger number of species, MDL morphology could predict foraging style across bat species and be included in future descriptions of foraging guilds.

Mammals can use a variety of physiological mechanisms to adapt to changes in their environment. Thyroid hormones (THs) are key modulators of growth and mediators of environmental conditions by regulating developmental processes and metabolism in animals. In recent years, advancements in non-invasive sampling have allowed monitoring of the fluctuations of THs and their metabolites in wild mammals. Triiodothyronine (T3) represents the major metabolite of THs excreted in feces so that it can be monitored in fecal samples. In this study, fecal samples collected during the hunting season from legally culled European roe deer (Capreolus capreolus; n = 160) were assayed to investigate the potential fluctuations of fecal TH metabolites (FTMs) in response to environmental (e.g., the temperature, local densities) and individual (e.g., sex, age, body, and nutritional conditions) variables. For this aim, we validated a TH enzyme immunoassay in the feces of roe deer. Our results show that FTMs can be successfully measured with satisfactory accuracy and precision. Extraction recovery (70%–120%), intra- and inter-day repeatability (<15%), linearity dilutions (80%–120%), and parallelism (<20%) were consistent with international guidelines. Environmental temperature (p < 0.001) showed a strong inverse correlation with FTM levels. THs can thus represent a reliable indicator in studying animals’ adaptative responses to environmental temperature changes, providing perspectives for the study of the impact of climate change on ungulates and mammals. Further analyses, comparing samples collected all year round, are needed to investigate the correlations of TH values versus the other investigated variables.

Over the past few decades, ocean hypoxia has been increasing due to human activities. Hypoxic stress, characterized by a reduced level of dissolved oxygen, is an escalating threat to marine ecosystems, with potentially devastating effects on the viability of endangered species such as the tri-spine horseshoe crab Tachypleus tridentatus. Even though this species is remarkably resilient to low oxygen levels, persistent hypoxia can negatively impact its population's survivability. The objective of this research was to examine the impact of short-term hypoxia on the behavior, gut microbiota, and metabolomics of juvenile T. tridentatus. First instar juvenile horseshoe crabs were subjected to hypoxic stress (2 mg O₂/L) for 14 days and then recovered for 7 days in an environment with normal dissolved oxygen. The findings demonstrated that short-term hypoxia reduced the rates of swimming and burrowing of horseshoe crabs, and induced the change of tissue metabolites and intestinal flora malfunction. Additionally, in the hypoxia groups on day 14, 86 distinct metabolites showed a trend of downregulation, while 29 metabolites showed an upregulation trend. Arginine biosynthesis; histidine metabolism; vitamin B6 metabolism; aminoacyl-tRNA biosynthesis; and alanine, aspartate, and glutamate metabolism were the top five metabolic KEGG pathways (p < 0.05) enriched with 8 metabolites. In conclusion, our results provided new insights related to the behaviors, microbiota, and metabolites involved in juvenile T. tridentatus exposed to short-term hypoxic conditions and confirmed that hypoxia impairs their behavioral and physiological status.

Identifying determinants of key phenotypic indicators driving animal population dynamics is fundamental to address measures aimed at mitigating human–wildlife interactions. In Mediterranean areas, summer drought reduces the availability of water and food resources for wild ungulates, potentially affecting body conditions. Since summer is a crucial period for the nursing/weaning of offspring, a seasonal bottleneck for ungulates would be expected to occur, especially in females. However, data on fluctuations of ungulate body mass during summer are scarce. We investigated the effects of summer progression, aridity, population density, and land use on body mass variation of adult/yearling wild boar (Sus scrofa) in a mixed forested-rural Mediterranean protected area, over a 16-year period (2007–2022). Data were gathered from individuals harvested during population control management actions, in summer. In contrast with our predictions, no consistent mass loss was observed throughout the summer. According to expectations, body mass was favored by decreasing aridity in the previous 3 months. In males, the positive effect of rainfall was mitigated by population density. Females experienced mass gain in non-arid years, mass loss in arid years, and no change in “average” years. The importance of edge habitats located at the interface between wooded and cultivated areas was confirmed by a positive relationship between female body mass and ecotone availability. The results indicate that, in a mixed forested-rural area, wild boar could maintain, or even increase, their body mass during summer. Moreover, findings emphasize the crucial role of rainfall in modulating a key driver of population dynamics for this ungulate in Mediterranean areas.

Greater thermal tolerance of invasive species benefits their survival and spread under extreme climate events, especially under global warming. Revealing the mechanisms underlying the interspecific differences in thermal tolerance between invasive and native species can help understand the invasion process and predict potential invaders. Here, we link the changes in global transcriptomics and antioxidant defense at multiple temperatures with the differences in thermal limits in the juveniles of a successful globally invasive turtle, Trachemys scripta elegans, and a native turtle in China, Mauremys reevesii. The two species show different thermal tolerances and have co-existed in habitats with the risk of overheating. The majority of the transcriptional response to thermal stress is conserved in the two turtle species, including protein folding or DNA damage responses activated under relatively moderate thermal stress and regulation of the cell cycle and apoptosis during severe thermal stress. Greater thermal tolerance of T. scripta elegans can be associated with a more stable global transcriptome during thermal stress, except for necessary stress responses, and a broader thermal range of continuous up-regulation of the core mechanisms promoting survival under thermal stress, mainly protein folding and negative regulation of apoptosis. Under extreme hot conditions, the opposite change trends of genes involved in survival mechanisms during thermal stress between invasive and native turtles can be due to differences in energy turnover. The present study provides insights into the mechanisms of physiological differences between invasive and native species given global transcriptional changes and helps understand successful invasion and predict potential invasive species.

Global climate change has heightened heat stress, threatening amphibian and reptile survival, including turtles. Although turtle species vary in heat tolerance, the molecular mechanisms behind these differences are not well understood. This study aimed to identify differentially expressed genes (DEGs) in response to heat stress (32°C) versus normal temperature (25°C) in eight tissues (brain, heart, intestine, liver, lung, muscle, spleen, and stomach) of two turtle species: Platysternon megacephalum (low heat tolerance) and Trachemys scripta elegans (high heat tolerance) using RNA-seq. The results revealed significant down-regulation of genes involved in energy and lipid metabolism in P. megacephalum, suggesting metabolic suppression under heat stress. Furthermore, the jumonji and AT-rich interaction domain containing 2 (JARID2) gene, which regulates cell proliferation and differentiation, was up-regulated in all tissues of P. megacephalum but down-regulated in all tissues of T. scripta elegans under heat stress. Pathway analysis revealed that protein processing in the endoplasmic reticulum was significantly enriched in brain, heart, lung, and muscle tissues of P. megacephalum, with BiP, CHOP, NEF, and HSPs significantly up-regulated in brain tissue, highlighting this pathway's impact on heat stress response. Seven hub genes were identified in the protein processing in the endoplasmic reticulum pathway in P. megacephalum. In contrast, T. scripta elegans showed a moderate response, with up-regulation of ribosomal genes in the brain to enhance protein synthesis and folding, while down-regulation of cell cycle genes in the intestine helped conserve energy for cellular repair. No significant pathways were found in other tissues of T. scripta elegans. These molecular responses in T. scripta elegans likely contribute to its better adaptation to heat stress. This study provides new insights into the molecular mechanisms of heat stress adaptation in turtles, offering valuable knowledge for understanding their ability to cope with future climate change.

Amphibians worldwide are declining due to various anthropogenic and environmental stressors. One of the most important threats is large-scale epidemics of chytridiomycosis, which is caused by Batrachochytrium dendrobatidis (Bd). Unlike in other continents, amphibian species in South Korea, such as Pelophylax nigromaculatus, are resistant to Bd, making it difficult to discern its detailed effects. This study determined the dynamics of Bd infection in P. nigromaculatus by integrating physiological, microbiological, and morphological data and applying state-of-the-art machine learning methodologies. Data are presented on Bd prevalence, body size, weight, and physiological stress responses, including corticosterone (CORT) levels and innate immune functions determined using bacterial killing assays and skin microbiome composition. Significant physiological differences between infected and non-infected animals were observed regarding elevated CORT levels and changes in bacterial killing capacity. Skin microbiome analysis indicated a subtle variation in the microbial composition, but the alpha and beta diversities did not significantly differ between infected and non-infected animals. To balance the intrinsic class imbalance of the dataset, several machine learning methods were coupled with different data-augmentation techniques. Using the Light Gradient Boosting Machine resulted in the best predictive performance when considering conditional generative adversarial networks-augmented datasets. Among the predictors, CORT level and bacterial killing ability were chosen for classifying the infection status. Machine learning can be used to complement the contrasting sensitivities of multi-level biomarkers due to differences in disease resistance or infection loads. This integrated approach may be essential for understanding the impacts of multiple threats to amphibians.

Colour morphs in polymorphic species are associated with a suite of heritable traits governed by distinct genetic loci, each corresponding to alternative fitness peaks. Hormonal pleiotropy has been proposed as a mechanism maintaining these morphs, though experimental evidence is lacking. In this study, we tested whether white and yellow morphs of the common wall lizard (Podarcis muralis) adopt alternative strategies shaped by the immuno-competence handicap hypotheses (ICHH). Specifically, we experimentally elevated plasma testosterone levels via transdermal administration and measured changes in (i) immune response using phytohaemagglutinin (PHA) swelling and (ii) the aggressive behaviour in response to a mirror test simulating territorial intrusion. We found that testosterone-induced immune suppression was stronger in white males, while aggression decreased in both morphs. Elevated testosterone eliminated the baseline differences between morphs in both immunity and aggression. These findings provide the first experimental support for morph-specific life-history strategies in common wall lizards, consistent with the ICHH: Yellow males prioritize aggression over immunity, while white males invest more in survival at the cost of competitiveness.

Variation patterns in gut microbial diversity among host species and populations offer valuable insights into the mechanisms underlying environment–host–microbiome interactions. However, the extent to which host phylogeny and geography drive these variation patterns in animal gut microbiota remains an open question. Amphibians and reptiles are important models to address this issue. Using 194 gut microbial samples of three amphibian and four reptile species inhabiting the Tarim Desert, we demonstrated phylosymbiosis in these animals, which was associated with heterogeneous selection and dispersal limitation. In interpopulation comparisons, Bufotes pewzowi and Teratoscincus przewalskii exhibited geography-dependent variations in their gut microbiota, particularly in relation to longitude and annual precipitation. These geographical patterns were linked to the heterogeneous selection of microbiota. Interestingly, the microbiota of B. pewzowi and T. przewalskii showed parallel variations with longitude, suggesting common selective pressures on amphibian and reptile microbiota. Finally, we found host genetic background did not account for the geographical pattern in their gut microbiota, even though it was also associated with geographical factors. This suggested environment–microbe interaction as a potential and independent ecological pathway mediating the associations between the environment and animals. Overall, these findings extend our understanding of variation patterns in gut microbiota and shed light on the underlying mechanisms.

The rumen microbiome plays a critical role in nutrient metabolism and adaptation of the yak (Bos grunniens), an import livestock animal of the Qinghai–Tibet Plateau renowned for their superior plant fiber degradation capacity. However, the microbiome among the different ecological niches within yak's rumen remains unelucidated. Through shotgun sequencing of rumen solid and liquid fractions from five yaks, we identified significant differences in the microbial communities and their genetic functions between the solid and liquid fractions. Solid fractions exhibited dominance by Ruminococcus, Succiniclasticum, and Aspergillus, while Prevotella, Paludibacter, Parabacteroides, and Bacteroides prevailed in liquid fractions. Comparative CAZyme profiling revealed solid fractions were significantly enriched in cellulose/hemicellulose-targeting enzymes (GH5, GH11, and CBM63), implicating their specialization in breaking down the fibrous grasses. In contrast, liquid fractions showed higher abundances of starch-degrading enzymes (GH13, CBM48) and host-glycan utilizers (GH92), suggesting roles in soluble nutrient extraction and host–microbe interactions. Comparative analysis of 574 metagenome-assembled genomes suggested that Methanomethylophilaceae_UBA71 and nitrate-respiring Ruminococcaceae_Firm-04 preferentially colonized in the solids, whereas propionate-producing Quinella and animal glycan-degrading Bacteroides were more prevalent in the liquids. Moreover, compared to Hu sheep, yak's rumen microbiome showed significantly enhanced utilization of plant polysaccharide capacity. Comparative analysis across 10 ruminant species further highlighted host phylogeny as a key driver of rumen microbiome variation. These findings advance our understanding of niche differentiation and functional specialization within the unique yak rumen ecosystem.

Female turtles are believed to select nesting sites that optimize conditions for egg development and allocate resources accordingly. Although relocating clutches to shaded hatcheries enhances hatchling survival, growth, and immune configuration, the impact of these unexpected environments on maternal investment remains uncertain. Herein, the effects of maternal (body size, as well as hematological and biochemical indicators) and environmental (sand temperature and moisture in both unshaded and shaded nests) variables on local inflammation after a challenge (as a proxy of immune function) and offspring size were evaluated using a split-clutch design. The association of maternal parameters with reproductive investment, as well as the relationship of incubation conditions to survival indicators (hatching and emergence success), was also evaluated. Hatchlings from shaded nests showed less intense inflammation and were heavier and longer compared to offspring from unshaded conditions. The incubation conditions influenced inflammation in hatchlings, as well as their body mass, straight carapace width, and survival. Maternal leukocytes differentially interacted with the environment to determine hatchling length. Maternal amylase and creatinine concentrations were related to clutch size and mass, respectively, while shading enhanced survival indicators. The results indicate that the incubation condition is the primary factor influencing hatchling phenotypes, both directly and indirectly by facilitating the expression of maternal effects. These findings suggest that while optimal environmental conditions and maternal quality do not synergize to enhance offspring fitness, environmental conditions can override the effects of maternal investment. This highlights the relevance of the incubation environment to guarantee hatchling phenotypes.

Increases in temperature associated with global warming have significant implications for organismal fitness. Thermal condition changes of inactive or dormant periods (such as winters) also have important effects on animals, particularly for ectotherms. Neglecting the potential consequences of winter warming can lead to biases in assessing the effect of climate change. The impacts of winter warming on ectotherms may be complex and multifaceted, possibly varying with geographic location including thermal ecological niche, altitude, and latitude. Therefore, we conducted field warming experiments (warmer climate vs. present climate) to investigate the effects of winter warming on the mass loss, body condition, physiological process, and survival capacity of Qinghai toad-headed lizards (Phrynocephalus vlangalii) at two contrasting altitudes (2600 vs. 3600 m) of the northern Qinghai–Xizang Plateau, China. The warming treatment reduced mass loss of the 2600-m-altitude lizard population, enhanced body condition, and increased overwintering survival rate after hibernation, while there was no significant effect on these indicators for the 3600-m-altitude lizard population with warming treatment. The two altitudinal populations showed different regulatory patterns of metabolic pathways in response to warming winters. Under simulated warming, the 2600-m-altitude lizard population mostly downregulated energy metabolism-related pathways (e.g., glycolysis, pyruvate metabolism, fatty acid degradation, TCA cycle, and oxidative phosphorylation) during hibernation. In contrast, under winter warming, the 3600-m-altitude lizard population primarily upregulated amino acid metabolism pathways (including serine and threonine metabolism; alanine, aspartate, and glutamate metabolism; cysteine and methionine metabolism; as well as histidine metabolism), which may be associated with cold stress adaptation. These findings contribute to our understanding of the adaptive effects of winter warming on reptiles and their physiological mechanisms, facilitating a better assessment of vulnerability to climate change.

Maintaining the body's water balance is crucial for function and survival in all animals. Humidity conditions vary between different habitats and greatly affect an animal's evaporative water loss (EWL). Species inhabiting arid regions have adaptions to minimize water loss, which those adapted to life in humid regions may lack. Therefore, the physiology of species from different habitats could respond differentially to acute exposure to dry conditions. We measured the EWL and resting metabolic rates (RMRs) of 12 Israeli squamate species, from either mesic or xeric habitats, spanning four orders of magnitude in size. We treated the animals to dry and humid air simulating natural conditions (vapor pressure deficits 3 and 1 kPa, respectively) at an ecologically relevant temperature of 25°C. EWL rates were higher in dry air, and the effect was stronger in mesic species. EWL of mesic species in humid air is similar to EWL of xeric species in dry air, indicating similar EWL when tested under settings that match each species’ natural conditions. In dry air, the RMR of small-bodied (<5 g) mesic species increased, whereas those of some small-bodied xeric species decreased. Small mesic species might be displaying stress from unnaturally dry conditions, whereas small xeric species possibly display an adaptation to minimize EWL by lowering RMR, thereby respiration rates. Physiological measurements are usually taken in dry air, and our results suggest previous experiments may contain a methodological bias. Future ecophysiological research needs to consider ambient humidity, by either varying experimental humidity to match natural conditions, or considering possible effects of humidity during analysis and interpretation of experiments and models.