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Phthalates have been shown to cause endocrine disruption in laboratory animals and are associated with altered development of the reproductive system in humans. Further, human have significant exposure to phthalates. However, little is known concerning the exposure of wildlife to phthalates. We report urinary phthalate metabolite concentrations from fifty juvenile alligators from three Florida lakes and a site in the Everglades. Urinary phthalate monoester concentrations varied widely among alligators from the different sites but also among alligators from the same site. Mono-2-ethylhexy phthalate and monobutyl phthalate were found in most samples of alligator urine with maximums of 35,700 ng/mL and 193 ng/mL, respectively. Monobenzyl phthalate was found in 5 alligators with a maximum of 66.7 ng/mL. Other monoesters were found in only one or two alligator urine samples. The wide variation within and among sites, in addition to the high levels of mEHP, mBP and mBzP, is consistent with exposure arising from the intermittent spraying of herbicide formulations to control invasive aquatic plants in Florida freshwater sites. Phthalate diesters are used as adjuvants in many of these formulations.

Sixteen monthly collections of adult male Gambusia holbrooki (eastern mosquitofish) were obtained from two lakes in central Florida, USA. Lake Woodruff and Lake Apopka are 36 miles apart, but differ in several environmental parameters. Compared with Lake Woodruff, Lake Apopka is warmer, more shallow in sampling areas (particularly during drought conditions; approximately 15-90 cm in Lake Apopka versus 60-120 cm in Lake Woodruff), more turbid, and more heavily contaminated with nutrients and industrial and agricultural chemicals. Here, we present detailed information on seasonal reproduction patterns in mosquitofish in their native range and compare patterns between fish from the two lakes. Male mosquitofish were reproductively active from spring through fall. Spermatogenesis, which is regulated in part by 11-ketotestosterone, ceased in October, and fish stored spermatozoa through the winter for immediate fertilization of offspring in the spring. Compared with Lake Woodruff, fish from Lake Apopka tended to be larger and have longer gonopodia and greater gonado- and hepato-somatic indices (GSI and HSI). High GSI in Apopka fish correlated with greater spermatid production, but fewer mature spermatozoa and either the same or lower sperm counts and sperm viability. Taken together, these observations suggest that differentiation of spermatids to spermatozoa is disrupted in Apopka fish, leading to reductions in fertility in some months. Delivery of sperm to females could also be affected in Apopka fish, which exhibit lower prevalence of efferent duct tissue in the testes during the summer.

Background The American alligator (Alligator mississippiensis) displays temperature-dependent sex determination (TSD), in which incubation temperature during embryonic development determines the sexual fate of the individual. However, the molecular mechanisms governing this process remain a mystery, including the influence of initial environmental temperature on the comprehensive gonadal gene expression patterns occurring during TSD. Results Our characterization of transcriptomes during alligator TSD allowed us to identify novel candidate genes involved in TSD initiation. High-throughput RNA sequencing (RNA-seq) was performed on gonads collected from A. mississippiensis embryos incubated at both a male and a female producing temperature (33.5 °C and 30 °C, respectively) in a time series during sexual development. RNA-seq yielded 375.2 million paired-end reads, which were mapped and assembled, and used to characterize differential gene expression. Changes in the transcriptome occurring as a function of both development and sexual differentiation were extensively profiled. Forty-one differentially expressed genes were detected in response to incubation at male producing temperature, and included genes such as Wnt signaling factor WNT11, histone demethylase KDM6B, and transcription factor C/EBPA. Furthermore, comparative analysis of development- and sex-dependent differential gene expression revealed 230 candidate genes involved in alligator sex determination and differentiation, and early details of the suspected male-fate commitment were profiled. We also discovered sexually dimorphic expression of uncharacterized ncRNAs and other novel elements, such as unique expression patterns of HEMGN and ARX. Twenty-five of the differentially expressed genes identified in our analysis were putative transcriptional regulators, among which were MYBL2, MYCL, and HOXC10, in addition to conventional sex differentiation genes such as SOX9, and FOXL2. Inferred gene regulatory network was constructed, and the gene-gene and temperature-gene interactions were predicted. Conclusions Gonadal global gene expression kinetics during sex determination has been extensively profiled for the first time in a TSD species. These findings provide insights into the genetic framework underlying TSD, and expand our current understanding of the developmental fate pathways during vertebrate sex determination. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-2396-9) contains supplementary material, which is available to authorized users.

Vertebrate genomes are mosaics of isochores, defined as long (>100 kb) regions with relatively homogeneous within-region base composition. Birds and mammals have more GC-rich isochores than amphibians and fish, and the GC-rich isochores of birds and mammals have been suggested to be an adaptation to homeothermy. If this hypothesis is correct, all poikilothermic (cold-blooded) vertebrates, including the nonavian reptiles, are expected to lack a GC-rich isochore structure. Previous studies using various methods to examine isochore structure in crocodilians, turtles, and squamates have led to different conclusions. We collected more than 6000 expressed sequence tags (ESTs) from the American alligator to overcome sample size limitations suggested to be the fundamental problem in the previous reptilian studies. The alligator ESTs were assembled and aligned with their human, mouse, chicken, and western clawed frog orthologs, resulting in 366 alignments. Analyses of third-codon-position GC content provided conclusive evidence that the poikilothermic alligator has GC-rich isochores, like homeothermic birds and mammals. We placed these results in a theoretical framework able to unify available models of isochore evolution. The data collected for this study allowed us to reject the models that explain the evolution of GC content using changes in body temperature associated with the transition from poikilothermy to homeothermy. Falsification of these models places fundamental constraints upon the plausible pathways for the evolution of isochores.

Several recent in situ studies have reported that domestic and mixed domestic/industrial sewage effluents contain one or more natural or anthropogenic estrogenic substances. Those studies examined caged or feral fish for the presence of the egg yolk precursor protein, vitellogenin (VTG), in the blood of male fish. We have previously reported that male, feral carp (Cyprinus carpio) obtained from the effluent channel of a major sewage treatment plant (STP) exhibited depressed serum testosterone (T) concentrations, as well as detectable levels of VTG. The present study examines male and female walleye (Stizostedion vitreum), a native species with a different life history and feeding habits, collected from the same Mississippi River locations below the St. Paul metropolitan STP. All male and female walleye collected from the effluent channel contained measurable levels of VTG in their blood. Males from that location also exhibited depressed serum T concentrations and elevated serum estradiol-17beta (E2) concentrations compared with males from the Snake River reference site. Males obtained from Mississippi River Navigational Pool #2 (MRP-2), 3-20 miles downstream of the STP also exhibited reduced serum T concentrations, but showed no alterations in E2 concentrations or the presence of VTG in the serum. Females collected at the STP site had greatly elevated serum E2 concentrations, but serum T concentrations were not different from females collected in the Snake River. Our results demonstrate that the St. Paul metropolitan STP continues to release an estrogenic effluent, capable of inducing VTG production and altering normal serum sex steroid concentrations in a commercially valuable, native fish, the walleye. Additional studies will be required to determine whether these observations portend long-term population level effects.

Concentrations of metals and selenium were examined in tissues of American alligators (Alligator mississippiensis) from three lakes in central Florida, in one of which alligators have exhibited reproductive or developmental defects. Our overall objective was to determine whether the levels of metals were sufficiently high to confound the association between chlorinated hydrocarbons, which are elevated in eggs and juvenile plasma, and reproductive impairment. The concentrations of all metals were relatively low compared to those reported for alligators from elsewhere in Florida and the southeastern United States, suggesting that reproductive impairment is not due to metals and that metals pose no health risk to the alligators. We also wanted to determine whether skin, biopsied tail muscle, or tail tip tissue, all easily collected from live alligators, could be used as surrogate measures of internal tissue loads. Concentrations of arsenic, cadmium, chromium, lead, manganese, mercury, and selenium in liver were highly correlated with at least one of the three biopsied tissues. Only tin showed no significant positive correlation. No single tissue gave a high prediction of liver levels for all metals, although skin gave the highest correlation for mercury, and tail muscle gave the best overall correlation for lead and cadmium.

Recent studies have reported a number of abnormalities in the hatchling and juvenile alligators of Lake Apopka, FL (USA). These abnormalities include modifications of plasma concentrations of sex steroids in males and females as well as abnormalities in gonadal morphology, gonadal enzyme activity, and steroidogenesis. Embryonic exposure to environmental contaminants in the eggs has been hypothesized to be the causal agent for these changes. However, posthatchling exposure can also contribute to changes in reproductive and endocrine functioning. We have detected serum concentrations of 16 of 18 organochlorine pesticides or metabolites (OCs) and 23 of 28 congener-specific polychlorinated biphenyls (PCBs) examined in juvenile alligators from Lake Apopka, Orange Lake, and Lake Woodruff National Wildlife Refuge. Lake Apopka juveniles had significantly elevated serum concentrations of p,p'-DDE, dieldrin, endrin, mirex, oxychlordane, SigmaDDTs, and SigmaPCBs compared to juveniles from the other lakes. Further, we observed no correlations between serum contaminant concentrations and sex steroid concentrations (estradiol-17beta and testosterone). However, serum testosterone was significantly lower in males from Lake Apopka and Orange Lake compared to Lake Woodruff NWR. We did not observe relationships between phallus size or other body parameters and serum contaminant levels. Phallus size was smaller in males from Lake Apopka even after adjustment for body size. We suggest that the observations previously reported for juvenile alligators-and observed again in this study-are apparently not associated with the current serum levels of the environmental contaminants we measured, but could be due to exposures during embryonic development to these or other pollutants. Future studies must determine if a causal relationship exists between the contaminants found in alligator eggs and abnormalities observed in the hatchlings and persisting in juveniles.