Previous studies have detected numerous organic contaminants and in vitro bioactivities in surface water from the South Platte River near Denver, Colorado, USA. To evaluate the temporal and spatial distribution of selected contaminants of emerging concern, water samples were collected throughout 2018 and 2019 at 11 sites within the S. Platte River and surrounding tributaries with varying proximities to a major wastewater treatment plant (WWTP). Water samples were analyzed for pharmaceuticals, pesticides, steroid hormones, and wastewater indicators and screened for in vitro biological activities. Multiplexed, in vitro assays that simultaneously screen for agonistic activity against 24 human nuclear receptors detected estrogen receptor (ER), peroxisome proliferator activated receptor-gamma (PPARγ), and glucocorticoid receptor (GR) bioactivities in water samples near the WWTP outflow. Targeted in vitro bioassays assessing ER, GR, and PPARγ agonism corroborated bioactivities for ER (up to 55 ± 9.7 ng/L 17β-estradiol equivalents) and GR (up to 156 ± 28 ng/L dexamethasone equivalents), while PPARγ activity was not confirmed. To evaluate the potential in vivo significance of the bioactive contaminants, sexually-mature fathead minnows were caged at six locations upstream and downstream of the WWTP for 5 days after which targeted gene expression analyses were performed. Significant up-regulation of male hepatic vitellogenin was observed at sites with corresponding in vitro ER activity. No site-related differences in GR-related transcript abundance were detected in female adipose or male livers, suggesting observed environmental concentrations of GR-active contaminants do not induce a detectable in vivo response. In line with the lack of detectable targeted in vitro PPARɣ activity, there were no significant effects on PPARɣ-related gene expression. Although the chemicals responsible for GR and PPAR-mediated bioactivities are unknown, results from the present study provide insights into the significance (or lack thereof) of these bioactivities relative to short-term in situ fish exposures.

Hypoxia is a major stressor in aquatic environments and it is frequently linked with excess nutrients resulting from sewage effluent discharges and agricultural runoff, which often also contain complex mixtures of chemicals. Despite this, interactions between hypoxia and chemical toxicity are poorly understood. We exposed male three-spined stickleback during the onset of sexual maturation to a model anti-androgen (flutamide; 250 μg/L) and a pesticide with anti-androgenic activity (linuron; 250 μg/L), under either 97% or 56% air saturation (AS). We assessed the effects of each chemical, alone and in combination with reduced oxygen concentration, by measuring the transcription of spiggin in the kidney, as a marker of androgen signalling, and 11 genes in the liver involved in some of the molecular pathways hypothesised to be affected by the exposures. Spiggin transcription was strongly inhibited by flutamide under both AS conditions. In contrast, for linuron, a strong inhibition of spiggin was observed under 97% AS, but this effect was supressed under reduced air saturation, likely due to interactions between the hypoxia inducible factor and the aryl hydrocarbon receptor (AhR) pathways. In the liver, hypoxia inducible factor 1α was induced following exposure to both flutamide and linuron, however this was independent of the level of air saturation. This work illustrates the potential for interactions between hypoxia and pollutants with endocrine or AhR agonist activity to occur, with implications for risk assessment and management.

Controversial glyphosate-based herbicides (GBHs) are the most frequently used herbicides globally. GBH residues in the wild, in animal and human food may expose non-target organisms to health risks, yet the developmental and cumulative effects of GBHs on physiology and reproduction remain poorly understood. We present the first long-term study on the effects of subtoxic GBH exposure (160 mg/kg) on multiple key physiological biomarkers (cellular oxidative status and neurotransmitters), gut microbiome, reproductive hormones, and reproduction in an avian model. We experimentally exposed in Japanese quail females and males (Coturnix japonica) to GBHs and respective controls from the age of 10 days–52 weeks. GBH exposure decreased hepatic activity of an intracellular antioxidant enzyme (catalase), independent of sex, but did not influence other intracellular oxidative stress biomarkers or neurotransmitter enzyme (acetylcholinesterase). GBH exposure altered overall gut microbiome composition, especially at a younger age and in females, and suppressed potentially beneficial microbes at an early age. Many of the microbial groups increased in frequency from 12 to 28 weeks under GBH exposure. GBH exposure decreased male testosterone levels both at sexual maturity and at 52 weeks of exposure, but did not clearly influence reproduction in either sex (maturation, testis size or egg production). Future studies are needed to characterize the effects on reproductive physiology in more detail. Our results suggest that cumulative GBH exposure may influence health and reproduction-related traits, which is important in predicting their effects on wild populations and global poultry industry.

Concern has increased regarding the adverse effects of polycyclic aromatic hydrocarbons (PAHs) on reproduction. However, limited information is available on the effects of PAHs in crustacean. In order to determine whether benzo[a]pyrene (B[a]P) could cause reproductive toxicity on the swimming crab Portunus trituberculatus, sexually mature female crabs were exposed to environmentally relevant concentrations of B[a]P (0, 0.1, 0.5 and 2.5 μg/L) for 10 days. B[a]P treatments resulted in high accumulation in ovary, and induced oxidative stress in a dose-dependent manner on ovary of crab. Furthermore, the haemolymph estradiol (E2) and testosterone (T) levels were significantly decreased. Histological investigation also revealed the reproductive toxicity caused by B[a]P. The results demonstrated that waterborne exposure to B[a]P caused oxidative damage and disrupted sex steroids in female crab P. trituberculatus, ultimately resulting in histological alternation.

Recently, bioaccumulation of dietary organic selenium (Se) in the ovaries and inhibition of reproduction in female aquatic animals have been reported. However, there is limited data on the subtle reproductive impacts of waterborne exposure to inorganic Se in fish. Here, zebrafish embryos (2 h post-fertilization) were exposed to solutions with environmentally relevant levels of Na₂SeO₃ with concentrations of 0 (control), 7.98 ± 0.31, 25.14 ± 0.15, and 79.60 ± 0.81 μg Se/L for 120 d until they reached sexual maturity. Female zebrafish were selected for reproductive toxicity assessment. In the early embryonic stage, whole-mount in situ hybridization of zebrafish embryos showed that waterborne Na₂SeO₃ exposure did not affect the observed location of vasa expression in primordial germ cells at 24, 48, and 72 h post-fertilization. Life-cycle exposure to 25.14 ± 0.15 and 79.60 ± 0.81 μg Se/L Na₂SeO₃ did not change the testosterone and 17β-estradiol contents in female zebrafish at the endpoint of exposure, but significantly reduced the proportion of early vitellogenic oocytes and mature oocytes. Follicle maturity retardation was accompanied by changes in transcriptional levels of the genes related to the hypothalamus-pituitary-gonad-liver (HPGL) axis. Transcriptional levels of cyp19a and lhr in the ovary were down-regulated, while the transcriptional level of fshr in the ovaries was up-regulated. In the 21-day cumulative spawning experiment, Na₂SeO₃ (25.14 ± 0.15 and 79.60 ± 0.81 μg Se/L) caused fewer eggs to be produced. Additionally, the malformation of zebrafish offspring significantly increased in the group exposed to 79.60 ± 0.81 μg Se/L. In conclusion, for the first time, this study shows that life-cycle exposure to environmentally relevant concentrations of waterborne Na₂SeO₃ significantly delays ovarian maturation and reduces the fertility of the female zebrafish.

Parabens are class of preservatives used in vast majority of commercial products, and a potential Endocrine Disrupting Chemical (EDC). The present study was undertaken to delineate the effects of n-butylparaben on F1 male progeny exposed maternally through gestation and lactation via subcutaneous route. The F0 dams were given subcutaneous injections of n-butylparaben from gestation day (GD) 6 to postnatal day (PND) 21 with doses of 10, 100, 1000 mg/kg Bw/day in corn oil. The F1 male rats were monitored for pubertal development and sexual maturation; these were sacrificed on PND 30, 45 and 75. On PND 75, these F1 male rats were subjected for fertility assessment with unexposed female rats.A delayed testicular descent at 100 and 1000 mg/kg Bw dose and delayed preputial separation at 10 mg/kg Bw dose was observed in exposed F1 male rats. Decreased sperm count, motility and Daily Sperm Production was observed at 100 mg/kg Bw dose at PND 75. Interestingly, the sperm transit time in the epididymis was accelerated at this dose. Significant perturbed testicular expression of steroid receptors (ERα and β, AR), INSL3 and StAR genes with increased T and LH levels indicates direct effect on spermatogenesis and steroidogenesis. These F1 generation adult rats were sub-fertile with increased (%) pre- and post-implantation loss at 100 and 1000 mg/kg Bw/day dose. This is the first report on n-butylparaben highlighting the involvement of testicular leydig cells with accelerated sperm transit time leading to reduced fertility in the maternally exposed F1 male rats through estrogenic/anti-androgenic action.

Perchlorate (ClO4−) contamination has been reported in ground and surface waters across North America. However, few studies have examined the effects of prolonged exposure to this thyroid hormone disrupting chemical, particularly at environmentally relevant concentrations in lower vertebrates, such as amphibians. The aim of this study was to examine the effects of a yearlong chronic exposure to ClO4− in adult male and female Western clawed frogs (Silurana tropicalis). Frogs were spawned and raised from fertilized embryo until sexual maturity in potassium perchlorate (KClO4)-treated water at different concentrations (0, 20, 53, and 107 μg/L). Developmental and reproductive indices – including adult morphology, androgen plasma levels, gonadal thyroid hormone- and sex steroid-related transcript levels, and sperm motility – were evaluated in male and female adult frogs. Female growth (e.g., body mass, snout-vent length, and hind limb length) was significantly reduced following chronic exposure to environmentally relevant concentrations of KClO4 resulting in females with morphometric indices similar to those of control males – indicating potential sex-specific sensitivities to KClO4. Changes to reproductive indices (i.e., plasma androgen levels, gonadal thyroid hormone- and sex steroid-related transcript levels, and sperm motility) were also observed in both sexes and suggest that KClO4 exposure may also have indirect secondary effects on the reproductive axes in male and female adult frogs. These effects were observed at concentrations at or below those reported in surface waters contaminated with ClO4− suggesting that this contaminant may have developmental and reproductive effects post-metamorphosis in natural amphibian populations.

Nitrate is a ubiquitous aquatic pollutant that is commonly associated with eutrophication and dead zones in estuaries around the world. At high concentrations nitrate is toxic to aquatic life but at environmental concentrations it has also been purported as an endocrine disruptor in fish. To investigate the potential for nitrate to cause endocrine disruption in fish, we conducted a lifecycle study with fathead minnows (Pimephales promelas) exposed to nitrate (0, 11.3, and 56.5 mg/L (total nitrate-nitrogen (NO3-N)) from <24 h post hatch to sexual maturity (209 days). Body mass, condition factor, gonadal somatic index (GSI), incidence of intersex, and vitellogenin induction were determined in mature male and female fish and plasma 11-keto testosterone (11-KT) was measured in males only. In nitrate-exposed males both 11-KT and vitellogenin were significantly induced when compared with controls. No significant differences occurred for body mass, condition factor, or GSI among males and intersex was not observed in any of the nitrate treatments. Nitrate-exposed females also had significant increases in vitellogenin compared to controls but no significant differences for mass, condition factor, or GSI were observed in nitrate exposed groups. Estradiol was used as a positive control for vitellogenin induction. Our findings suggest that environmentally relevant nitrate levels may disrupt steroid hormone synthesis and/or metabolism in male and female fish and may have implications for fish reproduction, watershed management, and regulation of nutrient pollution.

Pyrethroid pesticides were analysed in liver of striped dolphin (Stenella coeruleoalba) from the Alboran Sea (south of Spain, Mediterranean Sea). The occurrence and bioaccumulation of pyrethroid insecticides in marine mammal tissues from the northern hemisphere had never been determined before. Pyrethroids were detected in 87% of the specimens with a mean total concentration of 300 ng g-1 lw ± 932 (range 2.7–5200 ng g-1 lw). Permethrin and tetramethrin were the main contributors to the pyrethroid profiles, with enantiospecific accumulation for the first and isomer specific accumulation for the latter. Bioaccumulation of pyrethroids was unlike that of persistent organic pollutants (POPs), as pyrethroid concentrations were not correlated to the maturity stage of the specimens. Concentrations slightly increased from calves to juveniles, whereas juveniles presented similar concentrations to adults. Metabolization of pyrethroids after achieving sexual maturity might account for this pattern.

This study aimed to evaluate the adverse effects of the insecticide imidacloprid (IMI) on male spermatogenesis, steroidogenesis, and DNA damage in sexually mature and immature rats. Forty male rats (mature and immature) were equally divided into four groups: two mature and two immature groups. IMI groups of both ages were orally administered IMI in corn oil at a concentration of 1 mg/mL for kg BW/day, whereas their respective controls were orally administered corn oil only (1 mL/kg of body weight) daily for 65 days. On day 66, the rats were lightly anesthetized and then euthanized by cervical dislocation. Whole blood was collected for hemogram, serum for hormonal profile, semen for sperm profile, and testes for gene expression and histopathological, and immunohistochemical examinations. The obtained results revealed that both sexually mature and immature rats orally exposed to IMI showed serious abnormalities in sperm morphology and concentrations, with an imbalance of sexual hormones. There were increases in the level of serum 8-hydroxy-2′-deoxyguanosine and in the percentage of comet (tailed) sperm DNA in the IMI-treated groups. The results exhibited the upregulation of a DNA damage tolerance gene (8-oxoguanine glycosylase 1) and downregulation of the activity of steroidogenic genes (nuclear receptor subfamily 5, group A, member 1 and 3β-hydroxysteroid dehydrogenase). Immunohistochemical examination of the B-cell lymphoma 2-associated X apoptotic protein in testicular sections showed various degrees of apoptosis in the spermatogonial cells of the IMI-treated rats compared to the control groups. These damaging effects of IMI were more pronounced in the sexually mature rats than in the immature rats. In conclusion, despite using a low dose of IMI in the present study, there were noticeable harmful consequences on the reproductive system at different stages of sexual maturity in male rats.