Effect of perfluorooctanesulfonate exposure on steroid hormone levels and steroidogenic enzyme activities in juvenile Silurana tropicalis.

The impact of the perfluoro-chemical, perfluorooctanesulfonate (PFOS), on gonadal steroidogenesis during sexual differentiation in Silurana tropicalis was examined because of its ubiquity in the environment, bioaccumulative nature and potential to disturb endocrine activity. A partial life cycle study exposing S. tropicalis to varying concentrations of PFOS 0.06, 0.13, 0.25, 0.50 and 1.0 mg PFOS/L [nominal]) was conducted. Gonad and plasma samples were collected from juvenile control specimens and organisms exposed to PFOS from early embryo through 150 days post-metamorphosis. Gonad CYP17, aromatase and 5α-reductase activities were measured. Plasma estradiol, testosterone, dihydrotestosterone (DHT) and gonadal testosterone were measured in both males and females. Increased plasma DHT and gonadal testosterone were found in PFOS-treated juvenile male S. tropicalis compared to controls. Decreased plasma estradiol, but not testosterone, was detected in PFOS-treated female S. tropicalis compared to controls. Plasma DHT was not detected and an increase in gonadal testosterone was detected in PFOS-treated female frogs. Female S. tropicalis exposed to PFOS exhibited a concentration-related decrease in the mean aromatase activity, but not 5α-reductase. PFOS exposure in male frogs induced a concentration-related increase in 5α-reductase activity, but did not alter aromatase activity compared to control frogs. A concentration-related increase in CYP 17,20-lyase activity, but not 17-hydroxylase activity, was found in both female and male S. tropicalis exposed to PFOS.

Splenic immunotoxicity in developing cane toads (Rhinella marina) from Bermuda.

The impacts of contaminated sediment from 2 ponds in Bermuda on immune function in newly metamorphosed cane toads were examined. In the present study, a partial life-cycle experiment exposing Gosner stage 20 cane toad tadpoles to pond sediment and laboratory culture water through metamorphosis and into a juvenile state was performed. A basic immunology battery, including general necropsy, spleen somatic index, spleen white pulp content, splenocyte tissue density, and splenocyte viability, was conducted in newly metamorphosed Rhinella marina exposed to Bermuda freshwater sediment and baseline specimens collected from 2 separate populations in south Texas and south Florida, USA. Immune function was evaluated using a lymphocyte proliferation assay with subset specimens infected with Mycobacterium chelonae. In the Bermuda population exposed to pond sediment, splenocyte tissue density was markedly lower and lymphocyte proliferation substantially less relative to cohorts exposed to control sediment and to the North American populations. Considerable increases in spleen weight and liver and spleen lesions related to M. chelonae infection were recorded in challenged Bermuda R. marina compared with unchallenged specimens. Overall, immune function in Bermuda R. marina was compromised compared with North American mainland R. marina regardless of treatment but more dramatically in specimens exposed to Bermuda pond sediments. Environ Toxicol Chem 2016;35:2604-2612. © 2016 SETAC.

Application of endocrine disruptor screening program fish short-term reproduction assay: Reproduction and endocrine function in fathead minnow (Pimephales promelas) and killifish (Fundulus heteroclitus) exposed to Bermuda pond sediment.

A modified tier 1 Endocrine Disruptor Screening Program (EDSP) 21-d fish short-term reproduction assay (FSTRA) was used to evaluate the effects of sediment exposure from freshwater and brackish ponds in Bermuda on reproductive fecundity and endocrine function in fathead minnow (Pimephales promelas) and killifish (Fundulus heteroclitus). Reproductively active male and female fish were exposed to control sediment and sediment from 2 freshwater ponds (fathead minnow) and 2 marine ponds (killifish) contaminated with polyaromatic hydrocarbons and metals via flow-through exposure for 21 d. Reproductive fecundity was monitored daily. At termination, the status of the reproductive endocrine system was assessed by the gonadosomatic index, gonadal histology, plasma steroids (estrogen [E2], testosterone [T], and 11-ketotestosterone [11-KT]), steroidogenic enzymes (aromatase and combined 3ß/17ß -hydroxysteroid dehydrogenase [3ß/17ß-HSD]), and plasma vitellogenin (VTG). Decreased reproductive fecundity, lower male body weight, and altered endocrinological measures of reproductive status were observed in both species. Higher plasma T levels in female minnows and 11-KT levels in both male and female minnows and female killifish exposed to freshwater and brackish sediments, respectively. Decreased female E2 and VTG levels and gonadal cytochrome P19 (aromatase) activity were also found in sediment exposed females from both species. No effect on female 3ß/17ß-HSD activity was found in either species. The FSTRA provided a robust model capable of modification to evaluate reproductive effects of sediment exposure in fish.

Effects of multiple chemical, physical, and biological stressors on the incidence and types of abnormalities observed in Bermuda's cane toads (Rhinella marina).

The interactive effects of contaminants and ultraviolet light (UV)-exposure on the incidence and types of abnormalities observed were measured in newly metamorphosed cane toads (Rhinella marina) from four Bermuda ponds contaminated with petrochemicals and metals. Abnormalities were compared in toadlets that were field-collected, reared in predator exclusion cages, reared in laboratory microcosms exposed to control media or corresponding pond media, and reared in laboratory microcosms exposed to UV-light and control media or media from two ponds. Percent abnormal for field-collected, cage-reared, and microcosm-reared toadlets were equivalent per site and ranged between 14% and 63%. All treatments produced similar limb abnormalities but the percentage of hind versus forelimb defects was statistically greater only in field-collected toadlets. UV-exposed control media did not induce abnormalities in larvae exhibiting no maternal effect, and did not alter the types of abnormalities observed in larvae exhibiting a maternal or latent effect. Site media treatments without UV exposure induced significant cephalic and limb abnormalities, proved additive to the observed maternal/latent effect, and produced limb defects predominantly in forelimbs. Concurrent exposure to site media and UV-light induced similar types of abnormalities but a significantly higher percentage of hind limb abnormalities (68-89%) than exposure to site media alone (7-13%). Our results suggest that the types of abnormalities expressed were principally determined by direct and/or transgenerational contaminant exposure, but that UV-light exposure caused limb abnormalities to occur primarily in the hind limbs, mirroring field observations. Our field observations also suggest that ectromelia and brachydactyly in some field-collected specimens may be predator-induced.

Triclosan and thyroid-mediated metamorphosis in anurans: differentiating growth effects from thyroid-driven metamorphosis in Xenopus laevis.

In a previously reported study, we used a standard metamorphosis anuran model to assess potential effect of the antibacterial agent triclosan (TCS) on normal prometamorphic Xenopus laevis. Results indicated that environmentally relevant TCS concentrations did not alter the normal course of thyroid-mediated metamorphosis in this standard anuran model. However, to examine potential effects of TCS exposure during premetamorphosis and to distinguish between effects on metamorphosis and effects on growth, a longer term TCS exposure study was conducted. Standard Nieuwkoop and Faber (NF) stage 47 X. laevis larvae were exposed for 32 days (ca. NF stage 59-60) via flow-through to four different concentrations of TCS: < 0.2 (control), 0.8, 3.1, 12.5, or 50.0 µg TCS/l. Primary endpoints were survival, hind limb length, body length (whole; snout-to-vent), developmental stage, wet whole body weight, thyroid histology, plasma thyroid hormone (TH) concentrations, TH receptor beta (TRß), and type II and III deiodinase (DI-2 and DI-3) expression. Endpoints measured to evaluate effects on thyroid-mediated metamorphosis including developmental stage, thyroid histology, TRß expression, DI-2 and DI-3 expression, and thyroid gland 3,5,3',5'-tetraiodothyronine (T4) and plasma T4 and 3,5,3'-triiodothyronine (T3) levels were not affected by TCS exposure. However, increased larval growth based on whole body length (0.78, 12.5, and 50 µg TCS/l), snout-vent length (3.1 and 12.5 µg TCS/l), and whole body weight (0.8, 12.5, and 50.0 µg TCS/l) was observed following 32-day TCS exposure. These results indicated that TCS exposure during pre- and prometamorphosis increased larval growth but did not alter the normal course of metamorphosis in X. laevis. The increased growth associated with TCS exposure was not unexpected and is generally consistent with the presence of reduced bacterial stressors in culture.