Optimization, validation and application of a high-throughput 96-well elution protocol for the quantification of psychoactive substances in influent wastewater.

Wastewater-based epidemiology (WBE) is based on the analysis of human metabolic excretion products (biomarkers) of xenobiotics in wastewater, to gain information about various lifestyles and health aspects of a population in an evidence-based manner. Due to the complex wastewater matrix and trace level occurrence of human biomarkers in the sewage network, it is crucial to have sensitive analytical procedures available. Additionally, to improve the value of WBE as a complementary epidemiological source, there is increasing pressure on the analysis of more compounds, more locations and more samples. A high-throughput method based on 96-well Oasis MCX solid-phase extraction (SPE), requiring less influent wastewater (2 mL), was developed in accordance with the European Medicines Agency guidelines. Validation was successful for 28 parent drugs and metabolites of antidepressants, opioids and drugs of abuse. The selection of biomarkers and quantification limit was chosen to be relevant for WBE and was predominantly 10 ng/L or below. The final method was successfully applied to 24-h composite samples of October 2019 (n = 27), obtained from an urban wastewater treatment plant in Leuven (Belgium).

Evaluation of clonal relatedness of extended-spectrum beta-lactamase-producing Proteus mirabilis isolates by quantitative antibiogram and RAPD typing.

OBJECTIVE: To delineate, using two different typing systems, the clonal relatedness of 40 isolates of extended-spectrum beta-lactamase (ESbetaIa)-producing Proteus mirabilis obtained over a period of 7 years in six hospitals in the Paris area and two in Pas-de-Calais. METHODS: Random amplified polymorphic DNA (RAPD) polymerase chain reaction typing was applied by using three random primers on the ESbetaIa-producing P. mirabilis isolates and on isogenic Escherichia coli strains with or without plasmids encoding the representative resistance pattern transferred from P. mirabilis. Quantitative antibiogram typing, which was also applied to the P. mirabilis isolates, was used to define the euclidean distance between these strains. RESULTS: After having demonstrated that P. mirabilis plasmids did not influence chromosomal DNA amplification, we could classify the ESbetaIa-producing P. mirabilis isolates into 12 groups based on RAPD fingerprints. The same isolates were classified into 19 groups by quantitative antibiogram typing. Despite this difference in group numbers, general concordance between the typing systems was observed. This allowed us to show that the greater number of isolates in some hospitals belonged to a single strain and that single isolates obtained in different hospitals generally represented unique strains. CONCLUSIONS: A small number of ESbetaIa-producing P. mirabilis strains was isolated during 7 years in the eight medical centers studied, and the number of different strains identified suggested that inter-hospital transfer had not occurred.