Assessing relative sensitivity of marine and freshwater bivalves following exposure to copper: Application of classical and novel genotoxicological biomarkers.

Determination of relative sensitivity of biota following exposures to contaminants including metals is important for environmental protection. Copper (Cu), although biologically essential can be highly toxic to biota if present at higher concentrations in the natural environment. Given its ubiquitous presence within coastal and inland water bodies, we compared Cu-induced genotoxicity in two ecologically important mussel species, the freshwater Dreissena polymorpha (DP) and marine Mytilus galloprovincialis (MG), along with its tissue specific accumulation. Novel biomarker in terms of induction of gamma H2AX (γ-H2AX) foci, along with comet assay and induction of micronuclei (MN) were used to determine DNA damage response (DDR) in these two species following exposure to a range of Cu concentrations (18, 32, 56 µg L-1) for 10 days. Concentration-dependent increases in Cu concentration in gill tissue, as determined by Inductively Coupled Plasma Mass Spectrometry (ICP-MS), were paralleled by a greater degree of genotoxicity. An induction of γ-H2AX foci was present in all Cu exposure concentrations, proving this technique to be a sensitive and suitable biomarker of genotoxicity in bivalves. The multi-biomarker approach adopted here suggests firstly that in parallel with MG, which is widely used to assess the health of marine and coastal environment, DP is also suitable representative of inland water bodies, and that there is a similar mechanism of action for the induction of genotoxicity between the two species, following exposure to Cu. Secondly, for genotoxicity assessment a battery of responses could simultaneously be assessed in these two bivalve species. Finally, for adequate protection of the environment it is vital to adopt a multi-biomarker, multi-species approach to determine adverse biological effects to gain a holistic understanding of the real threat posed by contaminants to hydrosphere.

Assessing the impact of low level laser therapy (LLLT) on biological systems: a review.

PURPOSE: Low level laser therapy (LLLT) in the visible to near infrared spectral band (390-1100 nm) is absorption of laser light at the electronic level, without generation of heat. It may be applied in a wide range of treatments including wound healing, inflammation and pain reduction. Despite its potential beneficial impacts, the use of lasers for therapeutic purposes still remains controversial in mainstream medicine. Whilst taking into account the physical characteristics of different qualities of lasers, this review aims to provide a comprehensive account of the current literature available in the field pertaining to their potential impact at cellular and molecular levels elucidating mechanistic interactions in different mammalian models. The review also aims to focus on the integral approach of the optimal characteristics of LLLT that suit a biological system target to produce the beneficial effect at the cellular and molecular levels. METHODS: Recent research articles were reviewed that explored the interaction of lasers (coherent sources) and LEDs (incoherent sources) at the molecular and cellular levels. RESULTS: It is envisaged that underlying mechanisms of beneficial impact of lasers to patients involves biological processes at the cellular and molecular levels. The biological impact or effects of LLLT at the cellular and molecular level could include cellular viability, proliferation rate, as well as DNA integrity and the repair of damaged DNA. This review summarizes the available information in the literature pertaining to cellular and molecular effects of lasers. CONCLUSIONS: It is suggested that a change in approach is required to understand how to exploit the potential therapeutic modality of lasers whilst minimizing its possible detrimental effects.

Pharmaceutical Metabolism in Fish: Using a 3-D Hepatic In Vitro Model to Assess Clearance.

At high internal doses, pharmaceuticals have the potential for inducing biological/pharmacological effects in fish. One particular concern for the environment is their potential to bioaccumulate and reach pharmacological levels; the study of these implications for environmental risk assessment has therefore gained increasing attention. To avoid unnecessary testing on animals, in vitro methods for assessment of xenobiotic metabolism could aid in the ecotoxicological evaluation. Here we report the use of a 3-D in vitro liver organoid culture system (spheroids) derived from rainbow trout to measure the metabolism of seven pharmaceuticals using a substrate depletion assay. Of the pharmaceuticals tested, propranolol, diclofenac and phenylbutazone were metabolised by trout liver spheroids; atenolol, metoprolol, diazepam and carbamazepine were not. Substrate depletion kinetics data was used to estimate intrinsic hepatic clearance by this spheroid model, which was similar for diclofenac and approximately 5 fold higher for propranolol when compared to trout liver microsomal fraction (S9) data. These results suggest that liver spheroids could be used as a relevant and metabolically competent in vitro model with which to measure the biotransformation of pharmaceuticals in fish; and propranolol acts as a reproducible positive control.

Diamondoid naphthenic acids cause in vivo genetic damage in gills and haemocytes of marine mussels.

Diamondoids are polycyclic saturated hydrocarbons that possess a cage-like carbon skeleton approaching that of diamond. These 'nano-diamonds' are used in a range of industries including nanotechnologies and biomedicine. Diamondoids were thought to be highly resistant to degradation, but their presumed degradation acid products have now been found in oil sands process-affected waters (OSPW) and numerous crude oils. Recently, a diamondoid-related structure, 3-noradamantane carboxylic acid, was reported to cause genetic damage in trout hepatocytes under in vitro conditions. This particular compound has never been reported in the environment but led us to hypothesise that other more environmentally relevant diamondoid acids could also be genotoxic. We carried out in vivo exposures (3 days, semi-static) of marine mussels to two environmentally relevant diamondoid acids, 1-adamantane carboxylic acid and 3,5-dimethyladamantane carboxylic acid plus 3-noradamantane carboxylic acid with genotoxic damage assessed using the Comet assay. An initial screening test confirmed that these acids displayed varying degrees of genotoxicity to haemocytes (increased DNA damage above that of controls) when exposed in vivo to a concentration of 30 µmol L(-1). In a further test focused on 1-adamantane carboxylic acid with varying concentrations (0.6, 6 and 30 µmol L(-1)), significant (P < 0.05%) DNA damage was observed in different target cells (viz. gills and haemocytes) at 0.6 µmol L(-1). Such a level of induced genetic damage was similar to that observed following exposure to a known genotoxin, benzo(a)pyrene (exposure concentration, 0.8 µmol L(-1)). These findings may have implications for a range of worldwide industries including oil extraction, nanotechnology and biomedicine.

Emerging risks from ballast water treatment: the run-up to the International Ballast Water Management Convention.

Uptake and discharge of ballast water by ocean-going ships contribute to the worldwide spread of aquatic invasive species, with negative impacts on the environment, economies, and public health. The International Ballast Water Management Convention aims at a global answer. The agreed standards for ballast water discharge will require ballast water treatment. Systems based on various physical and/or chemical methods were developed for on-board installation and approved by the International Maritime Organization. Most common are combinations of high-performance filters with oxidizing chemicals or UV radiation. A well-known problem of oxidative water treatment is the formation of disinfection by-products, many of which show genotoxicity, carcinogenicity, or other long-term toxicity. In natural biota, genetic damages can affect reproductive success and ultimately impact biodiversity. The future exposure towards chemicals from ballast water treatment can only be estimated, based on land-based testing of treatment systems, mathematical models, and exposure scenarios. Systematic studies on the chemistry of oxidants in seawater are lacking, as are data about the background levels of disinfection by-products in the oceans and strategies for monitoring future developments. The international approval procedure of ballast water treatment systems compares the estimated exposure levels of individual substances with their experimental toxicity. While well established in many substance regulations, this approach is also criticised for its simplification, which may disregard critical aspects such as multiple exposures and long-term sub-lethal effects. Moreover, a truly holistic sustainability assessment would need to take into account factors beyond chemical hazards, e.g. energy consumption, air pollution or waste generation.

The effect of dietary organic chromium on specific growth rate, tissue chromium concentrations, enzyme activities and histology in common carp, Cyprinus carpio L.

A 63-day feeding trial was carried out to investigate the effect of three levels of Cr yeast (0.5, 1.0 and 2.0 mg Cr/kg) on the utilization of diets containing 38.5 % of maize starch or dextrin in common carp, Cyprinus carpio L. (initial mean body mass 14 ± 0.3 g) in an auto circulator system at 25 ± 0.5 °C. A two-way analysis of variance (ANOVA) showed that the final body mass (FBM), percentage mass gain (%MG), specific growth rate (SGR) and feed conversion ratio (FCR) were significantly (P < 0.05) affected by the two sources of variation (carbohydrate source and Cr level). In general, fish fed on a diet containing starch and fortified with 0.5 mg Cr/kg performed significantly higher FBM (47.23 g), %MG (225.11), SGR (1.91) and lower value of FCR (1.24) compared to fish fed on the other diets. Carp fed on 2.0 mg Cr/kg with maize starch and 1.0 mg Cr/kg with dextrin-based diet showed a significant reduction (P < 0.05) in whole body lipid content as confirmed by a two-way ANOVA. Fish fed on a maize starch-based diet supplemented with 0.5 and 1.0 mg Cr/kg recorded the highest activities for hexokinase enzyme. Glucose-6-phosphate dehydrogenase activity was neither affected by Cr concentration nor by dietary carbohydrate source. Fish fed on dextrin-based diets accumulated higher Cr in the whole tissue compared to fish fed on starch-based diets. Normal histological structures in the liver and gut tissues were observed in all groups. The present data clearly showed that dietary Cr yeast was safe in the fish diet at the levels tested.

The efficacy of chromium as a growth enhancer for mirror carp (Cyprinus carpio L): an integrated study using biochemical, genetic, and histological responses.

A growth trial was conducted on juvenile mirror carp (Cyprinus carpio L.) for 8 weeks to compare the efficacy of three chromium (Cr) compounds (Cr chloride, Cr picolinate, and Cr yeast) at a level 0.5 mg/kg as a potential growth enhancer. In addition, a high level of Cr (2.0 mg/kg) as Cr chloride has also been added in parallel for comparison. All Cr fortified diets at a level 0.5 mg/kg produced superior growth for carp compared to the control group and the group fed the high level of Cr chloride (2.0 mg/kg). Metabolic indicators measured included two of the key liver enzymes (hexokinase, HK) and (glucose-6-phosphate dehydrogenase, G6PD) activity. The results validated the positive effect of Cr at a level 0.5 mg/kg on enzyme activity and carbohydrate utilization producing significantly better growth performance for mirror carp. The study also included measurement of DNA strand breaks in the erythrocytes using the comet assay which revealed significantly (P < 0.05) increased DNA damage in fish fed on high level of Cr chloride (2.0 mg/kg) but the other treatments were not significantly different (P > 0.05) from the control groups. The concentration of Cr in the liver, gut, and whole fish tissues increased with increasing dietary Cr supplementation. Overall, Cr supplementation at a level 0.5 mg/kg from different sources may affect growth performance in carp by activation of some key liver enzymes (HK and G6PD).

Ecotoxicological applications and significance of the comet assay.

Application of the single-cell gel electrophoresis or comet assay has revolutionized the field of genetic ecotoxicology or eco-genotoxicology. It is a rapid, sensitive and relatively inexpensive method providing the opportunity to study DNA damage (including oxidative damage), repair and cell death (apoptosis) in different cell types without prior knowledge of karyotype and cell turnover rate. The assay has, however, often attracted criticism for its lack of ecotoxicological relevance. In addition, in contrast to genetic toxicology where rapid technical progress has been made to improve cell- and tissue-specific adoption of the assay, only limited advancement has been made to transfer the methodologies to ecotoxicological studies. While reviewing the recent information available in the literature and underscoring the importance of induced genetic damage in natural species, the aims of this article are to (i) highlight and judiciously analyse the ecotoxicological relevance of the assay; (ii) attempt to correlate the comet response with other relevant biological responses or biomarkers; (iii) identify the technical challenges and various factors affecting its application in order to make it reliable, reproducible and robust; (iv) critically compare the technical developments in genetic toxicology and genetic ecotoxicology and (v) evaluate the future developments with respect to applications of the assay. It is suggested that while complementing other ecotoxicological parameters and further improving the methodologies, the comet assay will continue to play an important role in genetic ecotoxicology to determine induced genetic damage, which has significant consequences for short- and long-term survival of the natural or wild species. Information obtained through integrated studies using simultaneous applications of multiple biomarkers on different wild organisms could also provide an holistic dimension of toxicological impact of environmental contaminants for the protection of human health.