Triclocarban (TCC), an antibacterial agent widely used in personal care products, can affect embryonic development. However, the specific molecular mechanism of TCC-induced embryonic developmental damage remains unclear. In this study, TCC exposure was found to increase the expression of tmbim4 gene in zebrafish embryos. The tmbim4 mutant embryos are more susceptible to TCC exposure than wild-type (WT) embryos, with tmbim4 overexpression reducing TCC-induced embryonic death in the former. Exposure of tmbim4 mutant larvae to 400 μg/L TCC substantially increased apoptosis in the hindbrain and eyes. RNA-sequencing of WT and tmbim4 mutant larvae indicated that knockout of the tmbim4 gene in zebrafish affects the autophagy pathway. Abnormalities in autophagy can increase apoptosis and TCC exposure caused abnormal accumulation of autophagosomes in the hindbrain of tmbim4 mutant zebrafish embryos. Pretreatment of TCC-exposed tmbim4 mutant zebrafish embryos with autophagosome formation inhibitors, substantially reduced the mortality of embryos and apoptosis levels. These results indicate that defects in the tmbim4 gene can reduce zebrafish embryo resistance to TCC. Additionally, apoptosis induced by abnormal accumulation of autophagosomes is involved in this process.

In order for Alberta's thick bitumen to be transported through pipelines, condensates are added creating a diluted bitumen (dilbit) mixture. Recent pipeline expansion projects have generated concern about potential dilbit spills on aquatic wildlife health. Studies have suggested that polycyclic aromatic compounds (PACs) are toxic to aquatic vertebrates and could potentially also interfere with their endocrine system. The research objectives of this study were to investigate the toxicity of dilbit to developing frog embryos and to identify the molecular mechanisms of action involved. Fertilized embryos of Western clawed frog (Silurana tropicalis) were exposed for 72 h to water accommodated fractions (WAF; 0.7–8.9 μg/L TPACs) and chemically-enhanced WAFs (CEWAF; 0.09–56.7 μg/L TPACs) of Access Western Blend (AWB) and Cold Lake Blend (CLB) dilbits. Both dilbit's CEWAFs significantly increased embryonic mortality and malformation incidence in the highest treatments tested, while WAF treatments led to no visible toxic effects. Increases of the cytochrome P450 1A (cyp1a) mRNA levels were observed for all WAF and CEWAF dilbit treatments suggesting that phase I detoxification is activated in the dilbit-exposed larvae. When exposed to PAC concentrations ranging from 0.09 to 8.9 μg/L, the frogs displayed no observable malformations, but expressed significant increases of cyp1a mRNA levels (2- to 25-fold; indicating a suitable biomarker of exposure); however, when concentrations were of 46.6 μg/L or higher, both malformed frog phenotype and induction of cyp1a mRNA level (>250-fold) were measured (indicating a suitable biomarker of response). The expression of several genes related to cellular detoxification and endocrine disruption were also measured, but were not significantly altered by the treatments. In sum, cyp1a mRNA level is a highly sensitive endpoint to measure subtle molecular changes induced by PAC exposure in the frog embryos and larvae, and data suggest that PAC concentration higher than 46 μg/L would be toxic to the developing S. tropicalis.

Ionizing radiation causes a variety of effects, including DNA damage associated to cancers. However, the effects in progeny from irradiated parents is not well documented. Using zebrafish as a model, we previously found that parental exposure to ionizing radiation is associated with effects in offspring, such as increased hatching rates, deformities, increased DNA damage and reactive oxygen species. Here, we assessed short (one month) and long term effects (one year) on gene expression in embryonic offspring (5.5 h post fertilization) from zebrafish exposed during gametogenesis to gamma radiation (8.7 or 53 mGy/h for 27 days, total dose 5.2 or 31 Gy) using mRNA sequencing. One month after exposure, a global change in gene expression was observed in offspring from the 53 mGy/h group, followed by embryonic death at late gastrula, whereas offspring from the 8.7 mGy/h group was unaffected. Interestingly, one year after exposure newly derived embryos from the 8.7 mGy/h group exhibited 2390 (67.7% downregulated) differentially expressed genes. Overlaps in differentially expressed genes and enriched biological pathways were evident between the 53 mGy/h group one month and 8.7 mGy/h one year after exposure, but were oppositely regulated. Pathways could be linked to effects in adults and offspring, such as DNA damage (via Atm signaling) and reproduction (via Gnrh signaling). Comparison with gene expression analysis in directly exposed embryos indicate transferrin a and cytochrome P450 2x6 as possible biomarkers for radiation response in zebrafish. Our results indicate latent effects following ionizing radiation exposure from the lower dose in parents that can be transmitted to offspring and warrants monitoring effects over subsequent generations.

Exposure of aquatic organisms to micro- and nano-sized plastic debris in their environment has become an alarming concern. Besides having a number of potentially harmful impacts for individual organisms, plastic particles can also influence the phenotype and performance of their offspring. We tested whether the sperm pre-fertilization exposure to nanoplastic particles could affect offspring survival, size, and swimming performance in the European whitefish Coregonus lavaretus. We exposed sperm of ten whitefish males to three concentrations (0, 100 and 10 000 pcs spermatozoa⁻¹) of 50 nm carboxyl-coated polystyrene spheres, recorded sperm motility parameters using computer assisted sperm analysis (CASA) and then fertilized the eggs of five females in all possible male-female combinations. Finally, we studied embryonic mortality, hatching time, size, and post-hatching swimming performance of the offspring. We found that highest concentration of plastic particles decreased sperm motility and offspring hatching time. Furthermore, sperm exposure to highest concentration of plastics reduced offspring body mass and impaired their swimming ability. This suggests that sperm pre-fertilization exposure to plastic pollution may decrease male fertilization potential and have important transgenerational impacts for offspring phenotype and performance. Our findings indicate that nanoplastics pollution may have significant ecological and evolutionary consequences in aquatic ecosystems.

With the recent ban of triclosan (TCS) and triclocarban (TCC) from some personal care products, many replacement antimicrobial compounds have been used. Yet the potential health risk and environmental impact of these replacement compounds are largely unknown. Here we investigated the toxicological effects of three commonly used replacement antimicrobials, benzalkonium chloride (BAC), benzethonium chloride (BEC), and chloroxylenol (CX) to two model organisms, the nematode C. elegans and zebrafish (Danio rerio), and compared them to the banned TCS and TCC. We found that these replacement compounds are not any safer than the banned antimicrobials. In the worm, at least one of the three, BAC, showed comparable toxicity to TCS from organismal to molecular levels, with toxic effects occurring at lower hundred μg/L to lower mg/L levels. In the fish, all three compounds at the tested concentration ranges (0.05–5 mg/L) showed toxicity effects to zebrafish embryos, indicated by hatching delay or inhibition, embryonic mortality, morphological malformations, and neurotoxicity. BAC was the most toxic among the three, with acute lethal toxicity occurring at environmentally relevant concentrations (hundreds of μg/L), which is comparable to the banned TCC. However, the toxicity effects of BAC and TCC occurred within different time windows, potentially suggesting different mechanisms of toxicity. CX was the only compound that induced a “body curvature” phenotype among the five compounds examined, suggesting a unique mode of toxic action for this compound. Furthermore, all five compounds except TCS induced neurotoxicity in fish larvae, indicated by alterations in secondary motoneuron axonal projections. Such neurotoxicity has been largely understudied for these antimicrobials in the past years and calls for further investigations in terms of its underlying mechanisms and ecological significance. These findings strongly indicate that scrutiny should be put on these replacement compounds before their introduction into massive use in personal care products.

This study aimed to explore the mechanism of perfluorooctanoic acid (PFOA) toxicity on the uterus and liver of mice during early pregnancy. Pregnant mice were given 0, 1, 5, 10, 20, and 40 mg/kg PFOA daily by gavage from gestational day (GD) 1–7 and sacrificed on GD 9. Subsequently, several toxicity parameters were evaluated, including the uterus and liver weights, liver and uterine indexes, histopathological changes of the liver and uterus, and levels of malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) in the liver. We also determined the expressions of FAS, FASL, Bax, Bcl-2, and Caspase-3 in decidual cells by immunohistochemistry and the TUNEL assay to detect apoptosis uterine cells. The results showed that PFOA increased the liver weights and reduced the uterus index in a dose-dependent manner. With increasing doses of PFOA, the levels of SOD and GSH-Px were significantly decreased, and MDA increased substantially in liver tissue. 20 mg/kg and 40 mg/kg of PFOA caused more substantial harm to the uterus, thus a higher probability for congestion and resorption. The expression of FAS, FASL, Bax, and Caspase-3 in decidual cells of the uterus in the PFOA treatment groups significantly increased in a dose-dependent manner. The expression of Bcl-2 was downregulated, decreasing the Bcl-2/Bax ratio. At gestation day 9, the control group had significantly fewer apoptotic cells in the uterus and shallower staining than the 40 mg/kg PFOA group. The findings of this study suggest that oxidative damage may be one of the mechanisms by which PFOA induces liver toxicity, and a subsequent increase in uterine cell apoptosis may cause embryo loss or damage.

Plants of the order Brassicaceae have evolved a chemical defence against herbivory: the glucosinolate-myrosinase system. Mechanical damage to plant tissues, such as grazing, initiates the production of phenethyl isothiocyanate (PEITC), a compound toxic to invertebrates. Mechanical damage caused during biofumigation and the harvesting and washing of watercress presents routes for PEITC release into waterbodies, such as the chalk stream spawning sites of brown trout (Salmo trutta). This laboratory study exposed developing S. trutta embryos to PEITC at concentrations of 0.01, 0.1 and 1 μg/L. S. trutta exposed to 1 μg/L PEITC during embryonic development resulted in 100% mortality after four dose days. Exposure to 0.1 μg/L PEITC resulted in an approximate fourfold increase in mortality relative to the controls, while exposure to 0.01 μg/L PEITC had a negligible effect on embryo mortality. Embryos exposed to 0.1 μg/L PEITC showed a significant delay in hatching and produced alevins with significantly shorter total lengths, lighter body weights and an approximate threefold increase in spinal deformities relative to those exposed to the controls and 0.01 μg/L PEITC. The results of a motor activity assay demonstrate that alevins exposed to PEITC showed a significant decrease in swimming activity compared with control animals during periods of illumination. The increased mortality, teratogenic effects and impaired behaviour in S. trutta following embryonic exposure to relatively low concentrations of PEITC highlight a need to accurately quantify and monitor environmental levels of PEITC.

Graphene has been used in several fields covering from electronics to biomedicine, especially exhibiting a widespread variety of promising biological and biomedical applications. In the past decade, the biomedical applications of graphene have attracted much interest. However, the effect of pristine graphene (pG) toxicity in aquatic vertebrates has not been fully studied. Thus, in this study, the toxicity of pG was experimentally evaluated using developing zebrafish embryos as in vivo model system. To determine this, 4-hpf embryos were exposed to different concentrations of pG (1, 5, 10, 15, 20, 25, 30, 35, 40, 45, and 50 μg/L) and different early life-stage parameters were observed at 24, 48, 72, and 96 hpf. Through embryogenesis, pG was observed to induce significant embryonic mortality, delayed hatching, heartbeat, several morphological defects, pericardial toxicity, and bradycardia. Yolk sac edema and pericardial edema were induced by pG in developing embryos. These outcomes would provide new insights into the adverse effects of pG on the developing embryonic cardiac defects in vertebrates and highlight the probable natural environment and health hazards of pG flakes.

Effects of radiating ostrich eggs before incubation on hatchability, growth performance, and some blood components of ostrich chicks were studied. 72 ostrich eggs were randomly distributed into four groups. The first group was kept without exposure to gamma radiation (control). The 2nd, 3rd, and 4th groups were exposed to 0.8, 1.6, and 3.2 Gy, respectively, of ⁶⁰Co gamma rays just before incubation. Total bacterial counts (×10³ CFU) were significantly increased with the increase in gamma radiation levels before incubation. Feed conversion was lower (P ≤ 0.01) in the control group than those irradiated. Embryonic mortality rate (%), hatchability (%) and chick weight at hatch, body weight, daily body weight gain, feed consumption, and all of hematology parameters studied were insignificantly fluctuated with the doses of gamma rays used. Serum concentrations of total protein, albumen, globulin, glucose, and triiodothyronin of the chicks from eggs of the control and the 1st group were significantly the highest. Chicks of the control group had significantly the lowest values of serum concentrations of uric acid, creatinine, triglyceride, total cholesterol, ALT, and AST when compared with the other groups. In conclusion, radiation of ostrich eggs before incubation with a dose of 0.8 Gy gamma has a stimulative effect upon the metabolism of hatched chicks.

BACKGROUND: Species that are at high levels of the food web have often been used as bioindicators to evaluate the presence of persistent contaminants in ecosystems. Most of these species are long-lived, so pollutant burdens may be integrated in some complex way over time. This makes them particularly sensitive to deleterious effects of Persistent Organic Pollutants (POPs). Birds have been suggested as useful organisms for monitoring pollutant levels. Traditionally such studies have been carried out with raptors such as osprey (Pandion haliaetus), peregrine falcon (Falco peregrinus), bald eagle (Haliaetus leucocephalus), etc. In this paper we present the results of a monitoring study conducted on two raptor species, osprey (Pandion haliaetus) and red kite (Milvus milvus), inhabiting a Mediterranean island (Menorca, Spain). These two species have different feeding habits; ospreys prey on fish and red kites feed on terrestrial species. This study constitutes a good opportunity to investigate if differences in feeding habits (aquatic vs. terrestrial) influences the contaminants pattern in two species inhabiting the same area. METHODS: The study was conducted in a non-destructive way, using only failed eggs, to avoid the damage of the population stability. Eggs were collected during the period 1994–2000. The contaminants examined were dichlorodiphenyltrichloroethanes (DDTs, including DDT and its main metabolite, DDE), polychlorinated biphenyls (PCBs), including ortho PCBs (PCBs with at least one Chlorine atom in the ortho position): #28, 52, 95, 101, 123+149, 118, 114, 153, 132+105, 138, 167, 156, 157, 180, 170, 189, 194; and non ortho PCBs (PCBs with no Chlorine atom in the ortho position): #77, 126, 169 and all the polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) with Chlorine atoms at the 2,3,7 and 8 position (2,3,7,8-substituted PCDDs and PCDFs). The analysis of organochlorine compounds was performed using a sample treatment based on a Solid Phase Matrix Dispersion procedure. Ortho PCBs and DDTs were determined by HRGC-µECD; non ortho PCBs and PCDD/Fs were determined by HRGC-HRMS. RESULTS AND DISCUSSION: The sum of the ortho PCB congeners analysed ranged from 0.94 to 15.03 µg/g wet weight (ww) for ospreys and from 1.0 to 11.2 µg/g ww for red kites. In both species, PCB congeners #153, #138 and #180 accounted about 75% to total ortho PCB concentrations. Regarding non ortho PCBs, for ospreys, concentrations ranged from 0.16 to 1.39 ng/g wet weight (ww) and for red kites from 0.12 to 0.51 ng/g ww, being congener #126 the most abundant. Concerning DDTs, concentration for ospreys ranged from 0.07 to 1.03 µg/g ww; and for red kites ranged from 0.90 to 2.10 µg/g ww, representing DDE more than 95% of the total DDTs, which proves a past use of DDT in the study area. Differences in contaminant levels between species are probably associated to feeding habits. The fish-eating species presents the highest PCB levels, whereas the terrestrial species exhibits the highest DDT levels. PCDD/Fs in ospreys were in the range 2.6–14.2 pg/g ww, while in red kites the range was slightly wider (22.2–43.2 pg/g ww), being PCDDs the major contributors in black kites. Ospreys had PCDDs similar to PCDF concentrations. PCDD/F profiles were mostly influenced by OCDD in both species. Non ortho PCBs were the major contributors to calculated Toxic Equivalent Quantity (TEQs) in both species. CONCLUSION: In both species studied, ortho-PCBs could represent a problem of concern since 57% of the eggs exhibited levels higher than 4 µg/g ww, reported as the level that could cause reduced hatchability, embryo mortality, and deformities in birds. RECOMMENDATIONS AND OUTLOOK: Results found in this study suggest that a more detailed study to clear up possible deleterious effects of PCBs on the bird populations studied here should be done.