Assessment of micropollutants toxicity by using a modified Saccharomyces cerevisiae model.

Environment can be affected by a variety of micropollutants. In this paper, we develop a system to assess the toxicity on an environmental sample, based on the expression of a nanoluciferase under the control of the STB5 promotor in a yeast. The STB5 gene encodes for a transcription factor involved in a pleiotropic drug resistance and in the oxidative stress response. The response of the modified yeast was assessed using 42 micropollutants belonging to different families (antibiotics, pain killers, hormones, plasticizers, pesticides, etc.). Among them, 26 induced an increase of the bioluminescence for concentration ranges from pg.L-1 to ng.L-1. Surprisingly, for concentrations higher than 100 ng.L-1, no response can be observed, suggesting that other mechanisms are involved when the stress increases. Analyzing the different responses obtained, we highlighted six nonmonotonic types of responses. The type of response seems to be independent of the properties of the compounds (polarity, toxicology, molecular weight) and of their family. In conclusion, we highlighted that a cellular response exists for very low exposition to environmental concentration of micropollutants and that it was necessary to explore the cellular mechanisms involved at very low concentration to provide a better risk assessment.

Comparison of Stir Bar Sorptive Extraction and Solid Phase Microextraction of Volatile and Semi-Volatile Metabolite Profile of Staphylococcus Aureus.

For the analysis of volatile bacterial compounds, solid phase microextraction (SPME) is currently the most widely used metabolite concentration technique. Recently, the potential of stir bar sorptive extraction (SBSE) for this use has been demonstrated. These two approaches were therefore used in combination with gas-chromatography coupled with mass-spectrometry (GC-MS) for the analysis of volatile and semi-volatile bacterial compounds produced by Staphylococcus aureus. In both cases, SPME and SBSE/headspace sorptive extraction (HSSE) enrichment was carried out in two coating phases. A whole analytical and statistical process was developed to differentiate the metabolites produced from the metabolites consumed. The results obtained with SBSE/HSSE and SPME were compared and showed the recovery of 90% of the compounds by SBSE/HSSE. In addition, we were able to detect the production of 12 volatile/semi-volatile compounds by S. aureus, six of which had never been reported before. The extraction by SBSE/HSSE showed higher concentration capacities and greater sensitivity than SPME concerning bacterial compounds, suggesting that this technique may therefore become the new preferred option for bacterial volatile and semi-volatile compound analysis.

Multiple stir bar sorptive extraction combined with gas chromatography-mass spectrometry analysis for a tentative identification of bacterial volatile and/or semi-volatile metabolites.

We propose a new approach combining the principles and advantages of stir bar sorptive extraction (SBSE) and headspace sorptive extraction (HSSE). Stir bars have so far never been used for the extraction of volatile/semi-volatile bacterial compounds. The effectiveness of two stir bars with polydimethylsiloxane (PDMS) or ethylene glycol/silicone (EGS) as sorbent was tested by performing sample extraction directly in gas chromatography (GC) vials containing bacterial cultures. Several combinations of desorption and extraction were tested at different growth times. When the extraction was carried out simultaneously with the EGS stir bar in headspace and the PDMS in the bacterial culture, the number of extracted compounds was significantly increased. Using both twisters increased the polarity range of the compounds found, and extraction at the end of the exponential phase of growth generated the best yields. This method was successfully applied to determine the production of 17 molecules by a strain of Staphylococcus aureus. In conclusion, this study paves the way for a new method for determining the volatile metabolite profile of bacteria, which can provide a promising innovative alternative in the identification of biomarkers.