Utilization of a Gender-Based Sorting Machine for Crustacean Selection in Bioconcentration Studies with the Freshwater Amphipod Hyalella azteca.

Bioconcentration factors (BCFs) are determined by fish flow-through tests performed according to Organisation for Economic Co-operation and Development test guideline 305. These are time-consuming and expensive and use a large number of animals. An alternative test design using the freshwater amphipod Hyalella azteca for bioconcentration studies has been recently developed and demonstrated a high potential. For bioconcentration studies using H. azteca, male amphipods are preferred compared with female organisms. Manual sexing of male adult amphipods is, however, time-consuming and requires care and skill. A new fully automatic sorting and dispensing machine for H. azteca based on image analysis has recently been developed by the company Life Science Methods. Nevertheless, an anesthesia step is necessary prior to the automatic selection. In the present study, we show that a single-pulse of 90 min of tricaine at the concentration of 1 g/L can be used and is recommended to select H. azteca males manually or automatically using the sorting machine. In the second part, we demonstrate that the machine has the ability to select, sort, and disperse the males of a culture batch of H. azteca as efficiently as manual procedures. In the last part of the study, BCFs of two organic substances were evaluated using the H. azteca bioconcentration test (HYBIT) protocol, with an anesthetizing step and robotic selection compared with manual selection without an anesthetizing step. The different BCF values obtained were in accordance with those indicated in the literature and showed that an anesthetizing step had no effect on the BCF values. Therefore, these data validated the interest in this sorting machine for selecting males to perform bioconcentrations studies with H. azteca. Environ Toxicol Chem 2023;42:1075-1084. © 2023 SETAC.

Scientific concepts and methods for moving persistence assessments into the 21st century.

The evaluation of a chemical substance's persistence is key to understanding its environmental fate, exposure concentration, and, ultimately, environmental risk. Traditional biodegradation test methods were developed many years ago for soluble, nonvolatile, single-constituent test substances, which do not represent the wide range of manufactured chemical substances. In addition, the Organisation for Economic Co-operation and Development (OECD) screening and simulation test methods do not fully reflect the environmental conditions into which substances are released and, therefore, estimates of chemical degradation half-lives can be very uncertain and may misrepresent real environmental processes. In this paper, we address the challenges and limitations facing current test methods and the scientific advances that are helping to both understand and provide solutions to them. Some of these advancements include the following: (1) robust methods that provide a deeper understanding of microbial composition, diversity, and abundance to ensure consistency and/or interpret variability between tests; (2) benchmarking tools and reference substances that aid in persistence evaluations through comparison against substances with well-quantified degradation profiles; (3) analytical methods that allow quantification for parent and metabolites at environmentally relevant concentrations, and inform on test substance bioavailability, biochemical pathways, rates of primary versus overall degradation, and rates of metabolite formation and decay; (4) modeling tools that predict the likelihood of microbial biotransformation, as well as biochemical pathways; and (5) modeling approaches that allow for derivation of more generally applicable biotransformation rate constants, by accounting for physical and/or chemical processes and test system design when evaluating test data. We also identify that, while such advancements could improve the certainty and accuracy of persistence assessments, the mechanisms and processes by which they are translated into regulatory practice and development of new OECD test guidelines need improving and accelerating. Where uncertainty remains, holistic weight of evidence approaches may be required to accurately assess the persistence of chemicals. Integr Environ Assess Manag 2022;18:1454-1487. © 2022 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Evaluating the ready biodegradability of two poorly water-soluble substances: comparative approach of bioavailability improvement methods (BIMs).

Difficulties encountered in estimating the biodegradation of poorly water-soluble substances are often linked to their limited bioavailability to microorganisms. Many original bioavailability improvement methods (BIMs) have been described, but no global approach was proposed for a standardized comparison of these. The latter would be a valuable tool as part of a wider strategy for evaluating poorly water-soluble substances. The purpose of this study was to define an evaluation strategy following the assessment of different BIMs adapted to poorly water-soluble substances with ready biodegradability tests. The study was performed with two poorly water-soluble chemicals-a solid, anthraquinone, and a liquid, isodecyl neopentanoate-and five BIMs were compared to the direct addition method (reference method), i.e., (i) ultrasonic dispersion, (ii) adsorption onto silica gel, (iii) dispersion using an emulsifier, (iv) dispersion with silicone oil, and (v) dispersion with emulsifier and silicone oil. A two-phase evaluation strategy of solid and liquid chemicals was developed involving the selection of the most relevant BIMs for enhancing the biodegradability of tested substances. A description is given of a BIM classification ratio (R BIM), which enables a comparison to be made between the different test chemical sample preparation methods used in the various tests. Thereby, using this comparison, the BIMs giving rise to the greatest biodegradability were ultrasonic dispersion and dispersion with silicone oil or with silicone oil and emulsifier for the tested solid chemical, adsorption onto silica gel, and ultrasonic dispersion for the liquid one.

High throughput and miniaturised systems for biodegradability assessments.

The society demands safer products with a better ecological profile. Regulatory criteria have been developed to prevent risks for human health and the environment, for example, within the framework of the European regulation REACH (Regulation (EC) No 1907, 2006). This has driven industry to consider the development of high throughput screening methodologies for assessing chemical biodegradability. These new screening methodologies must be scalable for miniaturisation, reproducible and as reliable as existing procedures for enhanced biodegradability assessment. Here, we evaluate two alternative systems that can be scaled for high throughput screening and conveniently miniaturised to limit costs in comparison with traditional testing. These systems are based on two dyes as follows: an invasive fluorescent dyes that serves as a cellular activity marker (a resazurin-like dye reagent) and a noninvasive fluorescent oxygen optosensor dye (an optical sensor). The advantages and limitations of these platforms for biodegradability assessment are presented. Our results confirm the feasibility of these systems for evaluating and screening chemicals for ready biodegradability. The optosensor is a miniaturised version of a component already used in traditional ready biodegradability testing, whereas the resazurin dye offers an interesting new screening mechanism for chemical concentrations greater than 10 mg/l that are not amenable to traditional closed bottle tests. The use of these approaches allows generalisation of high throughput screening methodologies to meet the need of developing new compounds with a favourable ecological profile and also assessment for regulatory purpose.