Testing androgen-induced immunosuppression: Environmental androgens as a model system for steroid-immune interaction.

It is well known that hormones influence and direct most facets of physiology; however, there is still contention regarding the directions of certain relationships, for example, between gonadal hormones and immunity. Among the many proposed relationships relating to gonadal-immune interactions, support for immunosuppressive effects of androgens remains prominent within physiological literature. Although ample study has been directed toward the immunosuppressive effects of androgens, considerable disagreement remains regarding their influence on immune function. In this study, we test the hypothesis that androgens inhibit immunocompetence in the American alligator (Alligator mississippiensis). Developing alligators were incubated at female-producing temperatures with a subset of individuals being exposed to 17-α-methyltestosterone (MT) before sexual determination. 17-α-methyltestosterone is a potent androgen, not aromatizable by crocodilians, that has been found to exert masculinizing effects in exposed crocodilian populations in vivo and in vitro. Additionally, a subset of animals was exposed to a novel antigen to quantify innate and acquired immune function. We recovered no significant differences in leukocyte ratios or proportions between groups and found no significant differences in innate immune function as measured by hemolysis-hemagglutination. However, we did find significant differences in acquired immune function, where masculinized individuals expressed greater antibody titers. Our findings reject the hypothesis that androgens suppress immune function; rather, androgens may be immunoenhancing to acquired humoral responses and neutral to innate humoral immunity in crocodilians.

The Spatial Ecology of Nuisance Crocodiles: Movement Patterns of Relocated American Crocodiles (Crocodylus acutus) in Guanacaste, Costa Rica.

Anthropogenic alterations of the environment have increased, highlighting the need for human-wildlife coexistence and conflict mitigation. Spatial ecology, and the use of passive satellite movement technology in particular, has been used to identify patterns in human-wildlife conflict as a function of shared resources that present potential for dangerous situations. Here, we aim to remotely identify patterns indicative of human-crocodile conflict in Guanacaste, Costa Rica by exploring site fidelity and diverse modes of movement (i.e., land and water) across space between nuisance (relocated) and non-nuisance (wild) crocodiles. Advanced satellite remote sensing technology provided near-constant movement data on individuals at the regional scale. Telonics Iridium SeaTrkr-4370-4 transmitters were used with modified crocodilian fitting. Results indicate that relocated crocodiles exhibited large-scale movements relative to wild crocodiles. Nuisance relocated crocodiles either returned to the area of nuisance or potentially attempted to in short time frames. The results presented here highlight the need for alternative management strategies that facilitate relocation efficacy.

The effect of androgen exposure on cerebral lateralization in the American alligator (Alligator mississippiensis).

The division of the brain manifests in lateralized physical behaviors, where specific tasks originate from one side of the body. Previous studies have shown that birds and reptiles mediate aggression in their right hemisphere and focus on opponents with their left eye. Degree of lateralization varies between sexes, likely due to androgen inhibition of lateralization in mammals, birds, and fish, but remains untested in herpetofauna. In this experiment, we investigated the effect of androgen exposure on cerebral lateralization in the American Alligator, Alligator mississippiensis. Alligator eggs were collected and incubated at female producing temperature with a subset dosed with methyltestosterone in ovo. Dosed hatchlings were randomly paired with control individuals and their interactions were recorded. The number of bites initiated by focus from each eye and the number of times an animal was bitten on each side of the body was recorded for each individual to elucidate cerebral lateralization in aggression. Control alligators had a significant bias towards left-eye bite initiation whereas androgen exposed alligators used both eyes indiscriminately. No significance was found in injury patterns. This study suggests that androgen exposure inhibits cerebral lateralization in alligator brains and corroborates right-hemisphere mediation of aggression, something previously unstudied in crocodilians.

The Inference of the Evolution of Immune Traits as Constrained by Phylogeny: Insight into the Immune System of the Basal Diapsid.

Among vertebrates, some of the most vulnerable taxa to emergent fungal pathogens are members of Reptilia. In light of the growing threat of emergent fungal pathogens affecting wildlife, it is important to broaden the current understanding of immune system function, development, and evolution. The homologous condition of a trait is necessary in order to study its evolution, as such, homology is necessary in the study of immunological evolution. Here, we explore the evolution of toll-like receptors (TLRs), a series of homologous receptors crucial to the initial immune response. The homologous condition of TLR genes provides a unique system in which to explore the evolution of the TLR; using a Reptilian phylogeny, we elucidate the immune condition of the basal diapsid. Our analysis revealed that the basal diapsid may have had an immune system that lacked two receptors: TLR 15, a receptor uniquely present in Reptilia, and TLR 13, a receptor important in the recognition of nucleic acid motifs. Additionally, our analysis showed multiple losses and convergences for various TLRs, likely attributed to redundancies in receptor function. Further exploration into the immune condition of extinct taxa may shed light on the evolution of the reptilian immune system.

Empirical and philosophical problems with the subspecies rank.

Species-level taxonomy derives from empirical sources (data and techniques) that assess the existence of spatiotemporal evolutionary lineages via various species "concepts." These concepts determine if observed lineages are independent given a particular methodology and ontology, which relates the metaphysical species concept to what "kind" of thing a species is in reality. Often, species concepts fail to link epistemology back to ontology. This lack of coherence is in part responsible for the persistence of the subspecies rank, which in modern usage often functions as a placeholder between the evolutionary events of divergence or collapse of incipient species. Thus, prospective events like lineages merging or diverging require information from unknowable future information. This is also conditioned on evidence that the lineage already has a detectably distinct evolutionary history. Ranking these lineages as subspecies can seem attractive given that many lineages do not exhibit intrinsic reproductive isolation. We argue that using subspecies is indefensible on philosophical and empirical grounds. Ontologically, the rank of subspecies is either identical to that of species or undefined in the context of evolutionary lineages representing spatiotemporally defined individuals. Some species concepts more inclined to consider subspecies, like the Biological Species Concept, are disconnected from evolutionary ontology and do not consider genealogy. Even if ontology is ignored, methods addressing reproductive isolation are often indirect and fail to capture the range of scenarios linking gene flow to species identity over space and time. The use of subspecies and reliance on reproductive isolation as a basis for an operational species concept can also conflict with ethical issues governing the protection of species. We provide a way forward for recognizing and naming species that links theoretical and operational species concepts regardless of the magnitude of reproductive isolation.

Latitudinal gradients in sexual dimorphism: Alternative hypotheses for variation in male traits.

Biological patterns across latitudinal gradients elucidate a number of striking natural clines from which numerous processes can be further explored. The trade-off between reproduction and somatic maintenance and growth represents a suite of life-history traits with variable energy allocation and potential latitudinal patterns. Specifically, male sexually dimorphic traits in female choice systems represent one such reproductive investment constrained by resource acquisition and subsequent allocation. Latitudinal variation in sexual dimorphism has been suggested although the relationship between dimorphic traits and latitude are conflicting. Here, we test alternative hypotheses regarding this pattern using two broadly distributed vertebrates exhibiting sexually dimorphic traits. We hypothesized that the exaggeration of dimorphic traits correlates with latitude, with males having exaggerated sexually dimorphic traits at either higher or lower latitudes. Results indicate that male sexually dimorphic traits are exaggerated at lower latitudes while relative gonopodium size in Poecilia latipinna was larger at higher latitudes. This pattern may be a result of lower latitude populations experiencing greater population densities and longer access to resources that could manifest in females more intensively selecting for higher quality males in lower latitudes. Experimental work should address this pattern and investigate mechanistic processes.

Testing the febrile response of snakes inoculated with Ophidiomyces ophidiicola, the causative agent of snake fungal disease.

Snake Fungal Disease (SFD) negatively impacts wild snake populations in the eastern United States and Europe. Ophidiomyces ophidiicola causes SFD and manifests clinically by the formation of heterophilic granulomas around the mouth and eyes, weight loss, impaired vision, and sometimes death. Field observations have documented early seasonal basking behaviors in severely infected snakes, potentially suggesting induction of a behavioral febrile response to combat the mycosis. This study tested the hypothesis that snakes inoculated with Ophidiomyces ophidiicola would seek elevated basking temperatures to control body temperature and behaviorally induce a febrile response. Eastern ribbon snakes (Thamnophis saurita, n = 29) were experimentally or sham inoculated with O. ophidiicola. Seven days after inoculation, snakes were tested on a thermal gradient and the internal body temperature and substrate temperature of each snake was recorded over time. Quantitative PCR was used when snakes arrived, during pre-inoculation, and post-inoculation to test snakes for the presence of O. ophidiicola. Some snakes arrived with O. ophidiicola and were subsequently inoculated, allowing for an assessment of secondary exposure effects. Snake thermoregulatory behavior was compared between 1) O. ophidiicola inoculated vs. sham inoculated treatments, 2) infected vs. disease negative groups, and 3) disease naïve vs. pre-exposed immune response categories. Neither internal nor substrate temperatures differed among initially prescribed, and qPCR recovered disease states, although infected snakes tended to reach a preferred body temperature faster than disease negative snakes. Snakes experiencing their first exposure (disease naïve) sought higher substrate temperatures than snakes experiencing their second exposure (pre-exposed). Here, we recover no evidence for behaviorally induced fever in snakes with SFD but do elucidate a febrile immune response associated with secondary exposure.

Use of continuous cranial shape variation in the identification of divergent crocodile species of the genus Mecistops.

The discovery of cryptic biodiversity has blossomed under the advancements of genetic techniques, but species identification via morphology remains crucial to effective conservation efforts. In this study, we tested the use of continuous cranial shape variation in distinguishing the two living species of Mecistops: the West African slender-snouted crocodile (M. cataphractus) and the Central African slender-snouted crocodile (M. leptorhynchus). Using a combination of geometric morphometric characters and ratios of linear measurements, we identified statistically significant variation in cranial bone and overall skull shape of mature individuals that corroborates existing molecular and discrete morphological evidence for two distinct, extant species within Mecistops. Specifically, variation in the shape of the nasal appears particularly diagnostic, while ratios involving metrics of snout length to snout width at the premaxillary notch offer distinguishing features easily measured in the field. Because of the complementary results and applications of the morphometric and cranial ratio analyses, we argue that both methodologies remain relevant to species identification. Moreover, we recommend continued cooperation between geneticists and morphologists in diagnosing species of conservation concern.

Microbial Assemblage Dynamics Within the American Alligator Nesting Ecosystem: a Comparative Approach Across Ecological Scales.

Understanding the ecological processes that shape species assemblage patterns is central to community ecology. The effects of ecological processes on assemblage patterns are scale-dependent. We used metabarcoding and shotgun sequencing to determine bacterial taxonomic and functional assemblage patterns among varying defined focal scales (micro-, meso-, and macroscale) within the American alligator (Alligator mississippiensis) nesting microbiome. We correlate bacterial assemblage patterns among eight nesting compartments within and proximal to alligator nests (micro-), across 18 nests (meso-), and between 4 geographic sampling sites (macro-), to determine which ecological processes may drive bacterial assemblage patterns within the nesting environment. Among all focal scales, bacterial taxonomic and functional richness (α-diversity) did not statistically differ. In contrast, bacterial assemblage structure (ß-diversity) was unique across all focal scales, whereas functional pathways were redundant within nests and across geographic sites. Considering these observed scale-based patterns, taxonomic bacterial composition may be governed by unique environmental filters and dispersal limitations relative to microbial functional attributes within the alligator nesting environment. These results advance pattern-process dynamics within the field of microbial community ecology and describe processes influencing the American alligator nest microbiome.

The evolution of crocodilian nesting ecology and behavior.

Crocodilians comprise an ancient and successful lineage of archosaurs that repeatedly raises questions on how they survived a mass extinction and remained relatively unchanged for ~100 million years. Was their success due to the change-resistant retention of a specific set of traits over time (phylogenetic conservatism) or due to flexible, generalist capabilities (e.g., catholic diets, phenotypic plasticity in behavior), or some combination of these? We examined the evolution of reproductive ecology and behavior of crocodilians within a phylogenetic perspective, using 14 traits for all 24 species to determine whether these traits were phylogenetically constrained versus (ecologically) convergent. Our analysis revealed that the ancestral crocodilian was a mound nester that exhibited both nest attendance and defense. Nesting mode exhibited 4-5 transformations from mound to hole nesting, a convergence of which habitat may have been a driving factor. Hole nesters were more likely to nest communally, but this association may be biased by scale. Although there were exceptions, mound nesters typically nested during the wet season and hole nesters during the dry season; this trait was relatively conserved, however. About two-thirds of species timed their nesting with the wet season, while the other third timed their hatching with the onset of the wet season. Nest attendance and defense were nearly ubiquitous and thus exhibited phylogenetic conservatism, but attendance lodging was diverse among species, showing multiple reversals between water and burrows. Collectively, our analysis reveals that reproductive trait evolution in crocodilians reflects phylogenetic constraint (nest attendance, nest defense), ecological convergence (seasonal timing of nesting, nest attendance lodging), or both (mode of nesting). Some traits (e.g., communal nesting and mode of nesting) were autocorrelated. Our analysis provides a framework for addressing hypotheses raised for why there has been trait convergence in reproductive ecology and behavior in crocodilians and why some traits remained phylogenetically conserved.

Critical thermal tolerance of invasion: Comparative niche breadth of two invasive lizards.

Understanding the evolution of thermal tolerance in ectotherms is particularly important given the current period of rapid change in the environment and thermal climate. Specifically, introduced species have the potential to evolve different thermal tolerances than native populations due to rapid evolution and novel selection pressures. Our study examined critical thermal tolerance in two introduced lizard species, the European wall lizard (Podarcis muralis) in Ohio and Kentucky, and the Mediterranean gecko (Hemidactylus turcicus) from Texas through Alabama. We tested the hypotheses that critical thermal maximum, minimum, and breadth varies among introduced populations of P. muralis and H. turcicus, and that critical thermal tolerance broadens when moving away from the introduction site, because dispersal across novel environments may remove dispersers with narrow thermal tolerances. We found that among P. muralis populations, CTmin and thermal breadth were significantly different. Specifically, when moving away from the introduction site, lizards exhibited increased cold tolerance and broader thermal breadth. Variability in thermal parameters were also lowest at the site closest to the introduction point in P. muralis. In contrast, H. turcicus had no significant differences in critical thermal minimum, maximum, or breadth among sites, or with respect to distance from introduction point. However, we did find little variability in thermal maximum, compared to greater variability in overall tolerance and critical thermal minimums. Thus, this study shows that selection on thermal tolerance and dispersal characteristics occur in novel climatic environments. Understanding how thermal tolerance changes over time can aid in predicting establishment and movement of introduced species, with applications for native species during a time of global climatic change.

Variability in snake skin microbial assemblages across spatial scales and disease states.

Understanding how biological patterns translate into functional processes across different scales is a central question in ecology. Within a spatial context, extent is used to describe the overall geographic area of a study, whereas grain describes the overall unit of observation. This study aimed to characterize the snake skin microbiota (grain) and to determine host-microbial assemblage-pathogen effects across spatial extents within the Southern United States. The causative agent of snake fungal disease, Ophidiomyces ophiodiicola, is a fungal pathogen threatening snake populations. We hypothesized that the skin microbial assemblage of snakes differs from its surrounding environment, by host species, spatial scale, season, and in the presence of O. ophiodiicola. We collected snake skin swabs, soil samples, and water samples across six states in the Southern United States (macroscale extent), four Tennessee ecoregions (mesoscale extent), and at multiple sites within each Tennessee ecoregion (microscale extent). These samples were subjected to DNA extraction and quantitative PCR to determine the presence/absence of O. ophiodiicola. High-throughput sequencing was also utilized to characterize the microbial communities. We concluded that the snake skin microbial assemblage was partially distinct from environmental microbial communities. Snake host species was strongly predictive of the skin microbiota at macro-, meso-, and microscale spatial extents; however, the effect was variable across geographic space and season. Lastly, the presence of the fungal pathogen O. ophiodiicola is predictive of skin microbial assemblages across macro- and meso-spatial extents, and particular bacterial taxa associate with O. ophiodiicola pathogen load. Our results highlight the importance of scale regarding wildlife host-pathogen-microbial assemblage interactions.

Early usage and meaning of evolvability.

Evolvability has become an enormously popular concept in evolutionary biology and in machine learning software architecture. While it is claimed that the term was coined in 1988 by Richard Dawkins, it was used as early as 1931 as a characteristic of life by John A. Thomson. We quote and review the earliest uses and definitions of evolvability in biological frameworks up until 1989, which are remarkably few. The meaning changed from simply the "ability to evolve" as a characteristic of life to various versions of including necessary variation to predict whether or not something could evolve to the rate and quality of that evolution. Or, meaning changed from the ability to evolve to the "quality" of the ability to evolve. Since then, evolvability has taken on many definitions as it has exploded in usage.

Host-specific phenotypic variation of a parasite co-introduced with invasive Burmese pythons.

Invasive Burmese pythons (Python bivittatus Kuhl, 1820) have introduced a lung parasite, Raillietiella orientalis, (Hett, 1915) from the python's native range in Southeast Asia to its introduced range in Florida, where parasite spillover from pythons to two families and eight genera of native snakes has occurred. Because these novel host species present a diversity of ecological and morphological traits, and because these parasites attach to their hosts with hooks located on their cephalothorax, we predicted that R. orientalis would exhibit substantial, host-associated phenotypic plasticity in cephalothorax shape. Indeed, geometric morphometric analyses of 39 parasites from five host species revealed significant variation among host taxa in R. orientalis cephalothorax shape. We observed differences associated with host ecology, where parasites from semi-aquatic and aquatic snakes exhibited the greatest morphological similarity. Morphological analyses of R. orientalis recovered from invasive pythons, native pit vipers, and terrestrial snakes each revealed distinct shapes. Our results suggest R. orientalis can exhibit significant differences in morphology based upon host species infected, and this plasticity may facilitate infection with this non-native parasite in a wide array of novel squamate host species.

Squamate areas of endemism in Cuba as determined by parsimony analysis of endemicity.

Cuba, the largest island in the Greater Antilles, hosts a high diversity of native squamate reptiles and is characterized by a complex geological history. The island has undergone repeated submergence and emergence, positioned on the dynamic border between the Caribbean and North American tectonic plates. Here, we infer current areas of endemism on Cuba based on squamate distributions using standard parsimony analysis of endemism under the "areas of endemism as individuals" hypothesis. We diagnose 29 areas of endemism, 14 of which are nested within other areas of endemism, from 52 squamate taxa. We suspect the current biotic pattern is a composite view of layered histories, and we summarize the geological history of the island to contemplate historical periods that left stronger marks on squamate distributions than others.

A salamander's top down effect on fungal communities in a detritivore ecosystem.

The soil decomposer community is a primary driver of carbon cycling in forest ecosystems. Understanding the processes that structure this community is critical to our understanding of the global carbon cycle. In North American forests, soil fungal communities are regulated by grazing soil invertebrates, which are in turn controlled by the predatory red-backed salamander (Plethodon cinereus). The presence of these soil invertebrate taxa is known to exert direct top-down control via selective grazing on saprotrophic fungi, with direct consequences for biogeochemical cycling in soil. We investigated whether the removal of P. cinereus would relieve top-down control on decomposer fungal communities in a tri-trophic mesocosm study. Fungal communities were characterized using metabarcoding and high-throughput DNA sequencing. The ß-diversity of fungal communities differed between salamander presence and absence treatments with a strong effect on saprotrophic fungal communities. We concluded that P. cinereus, a mesopredator in the detritivore food chain, exerts a prominent control on the composition and functional diversity of fungal communities in soil through a multi-trophic top-down process. Given their capacity to govern the compositions of soil invertebrates, the activity of these amphibians may be important for regulating ecosystem function and nutrient cycling in temperate forest systems.

Objectively measuring subjectively described traits: geographic variation in body shape and caudal coloration pattern within Vieja melanura (Teleostei: Cichlidae) / Midiendo objetivamente caracteres subjetivos: variación geográfica en la forma del cuerpo y patrones de coloración caudal de la especie Vieja melanura (Teleostei: Cichlidae)

AbstractVieja melanura is a Neotropical cichlid occurring in the Petén-lake district systems of Guatemala, as well as the Río Grijalva-Usumacinta basin, and other systems in Southern México, Belize, and Guatemala. A caudal stripe, extending forward from the caudal peduncle, is characteristic of this species. This stripe is sloped downward in nearly all individuals of V. melanura, but the degree of the slope is highly variable throughout its range. The slope and shape of the stripe has previously been used in diagnosing and differentiating between species of Vieja. The purpose of this study was to use objective methods to investigate morphological variation in the caudal stripe and body shape throughout the range of V. melanura. We studied geometric morphometric analyses of body shape and empirical measurements of the slope of the caudal stripe in 215 specimens ofV. melanura. We also used the mitochondrial cytochrome b marker to study population level patterns within V. melanura. Results from our analyses showed significant geographic variation in body shape and patterns of coloration with little mitochondrial phylogeographic structure. These patterns likely correspond to differences in riverine habitats throughout the species' distribution. In conclusion, these results can be used to inform other studies of color and shape variation as it applies to taxonomy and systematics.

ResumenViejamelanura es un cíclido neotropical (subfamilia Cichlinae; tribu Heroini) que se encuentra en los lagos del distrito de Péten en Guatemala, así como en la Cuenca del Río Grijalva-Usumacinta, y otros sistemas acuáticos en el sur de México, Belice y Guatemala. Una banda oscura que se extiende desde el pedúnculo caudal hacia la porción anterior del mismo es característica de esta especie. Esta banda está inclinada hacia abajo en casi todos los individuos de V. melanura, pero el grado de inclinación es altamente variable a lo largo del rango de distribución de la especie. La pendiente y forma de la banda se ha utilizado anteriormente como carácter diagnóstico entre especies del género Vieja. El propósito de este estudio fue investigar la variación morfológica en la forma del cuerpo y en los patrones de coloración de la banda caudal a lo largo del rango de distribución de V. melanura utilizando métodos objetivos. Analizamos la forma del cuerpo utilizando morfometría geométrica, medimos empíricamente la pendiente de la banda caudal en 215 especimenes. Finalmente evaluamos si existen patrones de estructura genética a nivel de poblaciones utilizando el marcador mitocondrial Citocromo b a lo largo del rango de distribución de V. melanura. Nuestros resultados muestran que la forma del cuerpo y los patrones de coloración de la banda caudal varian significantemente a lo largo del rango de distribución de V. melanura pero presenta poca estructura filogeografica. Estos patrones pueden ser explicados plausiblemente por diferencias en los hábitats riverinos presentes a lo largo del rango de distribución de la especie. En conclusión estos resultados se pueden utilizar como referencia en otros trabajos de taxonomía y sistemática que investiguen variación en la forma del cuerpo y patrones de coloracion en peces neotropicales.

Detection of a synthetic sex steroid in the American crocodile (Crocodylus acutus): Evidence for a novel environmental androgen.

Endocrine-disrupting contaminants (EDC's) are well known to alter sexual differentiation among vertebrates via estrogenic effects during development, particularly in organisms characterized by temperature-dependent sex determination. However, substances producing androgenic effects typically lack potency when tested in laboratory settings and are virtually unstudied in field settings. Here, we assay levels of a synthetic androgen, 17α-methyltestosterone (MT), in a heavily male-biased population of American crocodiles in the Tempisque River Basin of Costa Rica based on the recent hypothesis that this chemical is an EDC in developing crocodilian embryos. The presence of MT was documented in all field-collected samples of egg yolk and in plasma of all age classes in among population of crocodiles. Hatchlings exhibited higher plasma MT concentrations (102.1 ± 82.8 ng/mL) than juveniles (33.8 ± 51.5) and adults (25.9 ± 20.8 ng/mL). Among populations, crocodiles captured in the Tempisque River (62.9 ± 73.7 ng/mL) were higher in MT concentration than those from Tarcoles (13.3 ± 11.4 ng/mL) and negative controls (0.001 ± 0.0002 ng/mL). A mechanism for the bio-transport of MT and its subsequent effects is proposed.

Regional warming and the thermal regimes of American crocodile nests in the Tempisque Basin, Costa Rica.

Spatial variation in global climate change makes population-specific responses to this enigmatic threat pertinent on a regional scale. Organisms with temperature-dependent sex determination (TSD) potentially possess a unique physiological susceptibility that threatens population viability if rapid environmental effects on sex ratios render populations non-viable. A heavily male-biased sex ratio for hatchling American crocodiles of the Tempisque Basin, Costa Rica requires assessment of how nest temperature affects sex determination at this site, how females might compensate for these effects when creating nests, and how current patterns of climate change might alter future sex ratios and survival in hatchling cohorts. We demonstrate high within-nest variation in temperature but predict a female bias at hatching based on nest temperatures quantified here. Further, our data suggest that egg size and metabolic heating associated with this factor outweighs microhabitat parameters and depth in influencing nest thermal regimes. Finally, we document regional warming in the Tempisque Basin over the last 15 years and project that further heating over the next 15 years will not yield hatchling sex ratios as male biased as those currently found at this site. Thus, we find no support for nest temperature or climate change as likely explanations for male-biased American crocodile (Crocodylus acutus) sex ratios in the Tempisque Basin.

Methyltestosterone alters sex determination in the American alligator (Alligator mississippiensis).

Effects of xenobiotics can be organizational, permanently affecting anatomy during embryonic development, and/or activational, influencing transitory actions during adulthood. The organizational influence of endocrine-disrupting contaminants (EDC's) produces a wide variety of reproductive abnormalities among vertebrates that exhibit temperature-dependent sex determination (TSD). Typically, such influences result in subsequent activational malfunction, some of which are beneficial in aquaculture. For example, 17-αmethyltestosterone (MT), a synthetic androgen, is utilized in tilapia farming to bias sex ratio towards males because they are more profitable. A heavily male-biased hatchling sex ratio is reported from a crocodile population near one such tilapia operation in Guanacaste, Costa Rica. In this study we test the effects of MT on sexual differentiation in American alligators, which we used as a surrogate for all crocodilians. Experimentally, alligators were exposed to MT in ovo at standard ecotoxicological concentrations. Sexual differentiation was determined by examination of primary and secondary sex organs post hatching. We find that MT is capable of producing male embryos at temperatures known to produce females and demonstrate a dose-dependent gradient of masculinization. Embryonic exposure to MT results in hermaphroditic primary sex organs, delayed renal development and masculinization of the clitero-penis (CTP).