Comparison of the Zebrafish Embryo Toxicity Assay and the General and Behavioral Embryo Toxicity Assay as New Approach Methods for Chemical Screening.

The movement away from mammalian testing of potential toxicants and new chemical entities has primarily led to cell line testing and protein-based assays. However, these assays may not yet be sufficient to properly characterize the toxic potential of a chemical. The zebrafish embryo model is widely recognized as a potential new approach method for chemical testing that may provide a bridge between cell and protein-based assays and mammalian testing. The Zebrafish Embryo Toxicity (ZET) model is increasingly recognized as a valuable toxicity testing platform. The ZET assay focuses on the early stages of embryo development and is considered a more humane model compared to adult zebrafish testing. A complementary model has been developed that exposes larvae to toxicants at a later time point during development where body patterning has already been established. Here we compare the toxicity profiles of 20 compounds for this General and Behavioral Toxicity (GBT) assay to the ZET assay. The results show partially overlapping toxicity profiles along with unique information provided by each assay. It appears from this work that these two assays applied together can strengthen the use of zebrafish embryos/larvae as standard toxicity testing models.

An extract of the marine alga Alaria esculenta modulates α-synuclein folding and amyloid formation.

The conversion of α-synuclein from its natively unfolded and α-helical tetrameric forms to an amyloid conformation is central to the emergence of Parkinson's disease. Therefore, prevention of this conversion may offer an effective way of avoiding the onset of this disease or delaying its progress. At different concentrations, an aqueous extract from the edible winged kelp (Alaria esculenta), was shown to lower and to raise the melting point of α-synuclein. Size fractionation of the extract resulted in the separation of these distinct activities. The fraction below 5kDa decreased the melting point of α-synuclein, whereas the fraction above 10kDa raised the melting point. Both of these fractions were found to inhibit the formation of amyloid aggregates by α-synuclein, measured by thioflavin T dye-binding assays; this effect was further confirmed by transmission electron microscopy showing the inhibition of fibril formation. Circular dichroism analysis suggested that the incubation of α-synuclein under fibrillation conditions resulted in the loss of substantial native helical structure in the presence and absence of the fractions. It is therefore likely that the fractions inhibit fibrillation by interacting with the unfolded form of α-synuclein.

Rapid amyloid fibril formation by a winter flounder antifreeze protein requires specific interaction with ice.

A typically α-helical antifreeze protein (wflAFP-6) from winter flounder, Pseudopleuronectes americanus, forms amyloid fibrils during freezing. In this study, the effects of distinct components of the freezing process were examined. Freezing of wflAFP-6 in the presence of template ice was shown to be necessary for rapid conversion to an amyloid conformation. Neither subfreezing temperature nor phase change was sufficient. Thus, specific interaction with the ice surface was essential. The ice-induced formation of amyloid appeared to be unique to this helical antifreeze, it required high concentrations of protein and it occurred over a range of pH values. These results define a method for rapid formation of amyloid by wflAFP-6 on demand under physiological conditions.

Shewanella halifaxensis sp. nov., a novel obligately respiratory and denitrifying psychrophile.

Indigenous bacteria found in the sediment of the Emerald Basin (depth of 215 m, Atlantic Ocean) located offshore of Halifax Harbour (Nova Scotia, Canada) were previously found to be able to degrade the explosive compound hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). In the present study, a novel obligately respiratory, denitrifying and RDX-mineralizing bacterium, designated strain HAW-EB4(T), was isolated from the marine sediment. This bacterium utilized peptone, yeast extract, Casamino acids, esters (Tweens 20, 40 and 80), sugars (N-acetyl-D-glucosamine, ribose), several C2 and C3 acids (acetate, pyruvate, lactate, propionate) and amino acids (serine, proline) as sole carbon and energy sources. Aerobically grown cells (in marine broth 2216 at 10 degrees C) contained C(14 : 0) (6 %), iso-C(15 : 0) (12 %), C(16 : 0) (20 %), C(16 : 1)omega7 (37 %), C(18 : 1)omega7 (7 %) and C(20 : 5)omega3 (7 %) as major membrane fatty acids, and Q7 (28.1 %) and MK-7 (60.9 %) as dominant respiratory quinones, consistent with deep-sea species of Shewanella. The novel bacterium had a DNA G+C content of 45 mol% and showed similarity to Shewanella species in terms of 16S rRNA and gyrB gene sequences (93-99 and 67.3-88.4 % similarity, respectively), with Shewanella pealeana being the most closely related species. Genomic DNA-DNA hybridization between strain HAW-EB4T and S. pealeana revealed a level of relatedness of 17.9 %, lower than the 70 % species cut-off value, indicating that strain HAW-EB4T (= NCIMB 14093T = DSM 17350T) is the type strain of a novel species of Shewanella, for which the name Shewanella halifaxensis sp. nov. is proposed.