Assessment of poly(hexamethylenebicyanoguanide-hexamethylenediamine) hydrochloride-induced developmental neurotoxicity via oxidative stress mechanism: Integrative approaches with neuronal cells and zebrafish.

Poly(hexamethylenebicyanoguanide-hexamethylenediamine) hydrochloride (PHMB) is a biocide with a broad spectrum of antibacterial activity. Its use as a disinfectant and preservative in consumer products results in human exposure to PHMB. Toxicity studies on PHMB mainly focus on systemic toxicity or skin irritation; however, its effects on developmental neurotoxicity (DNT) and the underlying mechanisms are poorly understood. In this study, the DNT effects of PHMB were evaluated using IMR-32 and SH-SY5Y cell lines and zebrafish. In both cell lines, PHMB concentrations ≥ 10 µM reduced neurite outgrowth, and cytotoxicity was observed at concentrations up to 40 µM. PHMB regulated expression of neurodevelopmental genes and induced reactive oxygen species (ROS) production and mitochondrial dysfunction. Treatment with N-acetylcysteine reversed the toxic effects of PHMB. Toxicity tests on zebrafish embryos showed that PHMB reduced viability and heart rate and caused irregular hatching. PHMB concentrations of 1-4 µM reduced the width of the brain and spinal cord of transgenic zebrafish and attenuated myelination processes. Furthermore, PHMB modulated expression of neurodevelopmental genes in zebrafish and induced ROS accumulation. These results suggested that PHMB exerted DNT effects in vitro and in vivo through a ROS-dependent mechanism, highlighting the risk of PHMB exposure.

Ecological Risk Assessment of Amoxicillin, Enrofloxacin, and Neomycin: Are Their Current Levels in the Freshwater Environment Safe?

Veterinary pharmaceuticals may cause unexpected adverse effects on non-target aquatic species. While these pharmaceuticals were previously identified as priority compounds in ambient water, their ecological risks are relatively unknown. In this study, a series of chronic toxicity tests were conducted for these pharmaceuticals using algae, two cladocerans, and a fish. After a 21-d exposure to amoxicillin, enrofloxacin, and neomycin, no observed effect concentration (NOEC) for the reproduction of Daphnia magna was detected at 27.2, 3.3, and 0.15 mg/L, respectively. For the survival of juvenile Oryzias latipes following the 40-d exposure, NOEC was found at 21.8, 3.2, and 0.87 mg/L, respectively. Based on the results of the chronic toxicity tests and those reported in the literature, predicted no-effect concentrations (PNECs) were determined at 0.078, 4.9, and 3.0 µg/L for amoxicillin, enrofloxacin, and neomycin, respectively. Their hazard quotients (HQs) were less than 1 at their average levels of occurrence in ambient freshwater. However, HQs based on the maximum detected levels of amoxicillin and enrofloxacin were determined at 21.2 and 6.1, respectively, suggesting potential ecological risks. As the potential ecological risks of these veterinary pharmaceuticals at heavily contaminated sites cannot be ignored, hotspot delineation and its management are required.

Comparative assessment of the adverse outcome of wastewater effluents by integrating oxidative stress and histopathological alterations in endemic fish.

This study evaluated the adverse effect of wastewater effluents on three fish species, Carassius auratus, Zacco platypus, and Zacco koreanus, collected in the Eungcheon, Mihocheon, and Busocheon streams, respectively. Fish gills, liver, and kidneys from the mixing zone (MZ) and sites upstream (US) and downstream (DS) of the MZ were analyzed for oxidative stress responses and histology. Catalase and glutathione S-transferase activity was significantly higher at MZ and DS than from US (p<0.05), indicating induction of antioxidant defense mechanisms. Additionally, degree of tissue changes (DTC) indicated highest histopathological alteration in MZ, followed by DS and US. Integrated biomarker response (IBR) for oxidative stress and histopathological alterations showed higher values consistently for Z. koreanus than other two species. Water temperature, EC, and TN levels seemed to be responsible for the observed biomarker responses. These findings indicate that thermal hot spring effluent discharged into Busocheon steam induced the most significant impact on the cool water species (Z. koreanus). Overall, this study suggests that the IBR index is a very useful tool for monitoring in situ adverse effects of wastewater effluents on fish, particularly for histopathological alterations representing prolonged impact.

In situ impact assessment of wastewater effluents by integrating multi-level biomarker responses in the pale chub (Zacco platypus).

The integration of biomarker responses ranging from the molecular to the individual level is of great interest for measuring the toxic effects of hazardous chemicals or effluent mixtures on aquatic organisms. This study evaluated the effects of wastewater treatment plant (WWTP) effluents on the freshwater pale chub Zacco platypus by using multi-level biomarker responses at molecular [mRNA expression of catalase (CAT), superoxide dismutase (SOD), glutathione S-transferase (GST), and metallothionein (MT)], biochemical (enzyme activities of CAT, SOD, GST, and concentration of MT), and physiological [condition factor (CF) and liver somatic index (LSI)] levels. The mRNA expression levels of GST and MT in Z. platypus from a site downstream of a WWTP significantly increased by 2.2- and 4.5-fold (p<0.05) when compared with those from an upstream site. However, the enzyme activities of CAT, SOD, and GST in fish from the downstream site significantly decreased by 43%, 98%, and 13%, respectively (p<0.05), except for an increase in MT concentration (41%). In addition, a significant increase in LSI (46%) was observed in Z. platypus from the downstream site (p<0.05). Concentrations of Cu, Zn, Cd, and Pb in the liver of Z. platypus were higher (530%, 353%, 800%, and 2,200%, respectively) in fish from a downstream site than in fish from an upstream location, and several multi-level biomarker responses were significantly correlated with the accumulated metals in Z. platypus (p<0.05). Integrated biomarker responses at molecular, biochemical, and physiological levels (multi-level IBR) were much higher (about 4-fold) at the downstream site than at the upstream site. This study suggests that the multi-level IBR approach is very useful for quantifying in situ adverse effects of WWTP effluents.

Integrative assessment of biomarker responses in pale chub (Zacco platypus) exposed to copper and benzo[a]pyrene.

Multi-level biomarker responses (molecular/biochemical and histological/physiological levels) were studied to assess the sublethal toxicities of copper (Cu: 1.25, 5, and 20µg/L) and benzo[a]pyrene (BaP: 0.5, 5, and 50µg/L) induced in the freshwater pale chub Zacco platypus. Except for the kidney tissues when exposed to 20µg Cu/L, no significant differences were observed at the histological or physiological levels among the treatment groups. However, various molecular and biochemical responses were observed in Z. platypus, and these responses primarily depended on exposure time. Upon Cu exposure, both DNA single-strand breaks (COMET) and metallothionein (MT) concentration significantly increased after 4 days, whereas there were no significant changes after 14 days of exposure. Both 4 and 14 days of BaP exposure induced significant increases in COMET and 7-ethoxyresorufin-O-deethylase (EROD) activity, but there was no significant difference between them. Additionally, both Cu and BaP induced acetylcholinesterase (AChE) activity only after 14 days of exposure. The current findings demonstrate that the differences in the responses of MT and EROD are associated with each chemical's particular mode of action. Biomarker responses at the molecular and biochemical levels were quantized in terms of the integrated biomarker response (IBR) index to compare the toxicities of Cu and BaP. The IBR values were well correlated with the concentrations of Cu and BaP, and the correlations were enhanced at 4 days of exposure (r(2)=0.849 and 0.945, respectively) compared with 14 days (r(2)=0. 412 and 0.634, respectively). These results suggest that the IBR index may be a useful tool for the integrative quantification of the molecular and biochemical biomarker responses in a short-term exposure to Cu and BaP.

Integrated assessment of biomarker responses in common carp (Cyprinus carpio) exposed to perfluorinated organic compounds.

In this study, the toxicological effects of perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) toward the common carp, Cyprinus carpio, were evaluated by assessing the responses of five biomarkers, including DNA single-strand breaks (COMET), vitellogenin (VTG) concentration, and the activities of 7-ethoxyresorufin-O-deethylase (EROD), acetylcholinesterase (AChE) and catalase (CAT). Upon PFOA exposure, both the VTG concentration and CAT activity were significantly increased, while there was a negligible change in the responses of other biomarkers when compared to the control. Upon PFOS exposure, a significant increase in the DNA single-strand breaks was observed, while the responses of other biomarkers were not significantly altered when compared to the control. Standardized scores of biomarker responses were visualized using star plots and computed as the integrated biomarker response (IBR). As expected from the different biomarker responses, PFOA and PFOS showed totally different patterns of star plots. Additionally, the IBR values were well correlated with the logarithmic concentrations of PFOA and PFOS (R(2)=0.9434 and 0.9511, respectively). These results suggest that the IBR might be a useful tool for quantification of various biomarker responses induced by toxic chemicals.