A workflow for the detection of antibiotic residues, measurement of water chemistry and preservation of hospital sink drain samples for metagenomic sequencing

Background: Hospital sinks are environmental reservoirs that harbour healthcareassociated (HCA) pathogens. Selective pressures in sink environments, such as antibiotic residues, nutrient waste and hardness ions, may promote antibiotic resistance gene (ARG) exchange between bacteria. However, cheap and accurate sampling methods to characterize these factors are lacking. Aims: To validate a workflow to detect antibiotic residues and evaluate water chemistry using dipsticks. Secondarily, to validate boric acid to preserve the taxonomic and ARG ('resistome') composition of sink trap samples for metagenomic sequencing. Methods: Antibiotic residue dipsticks were validated against serial dilutions of ampicillin, doxycycline, sulfamethoxazole and ciprofloxacin, and water chemistry dipsticks against serial dilutions of chemical calibration standards. Sink trap aspirates were used for a 'realworld' pilot evaluation of dipsticks. To assess boric acid as a preservative of microbial diversity, the impact of incubation with and without boric acid at -22 degrees C on metagenomic sequencing outputs was evaluated at Day 2 and Day 5 compared with baseline (Day 0). Findings: The limits of detection for each antibiotic were: 3 mu g/L (ampicillin), 10 mu g/L (doxycycline), 20 mu g/L (sulfamethoxazole) and 8 mu g/L (ciprofloxacin). The best performing water chemistry dipstick correctly characterized 34/40 (85%) standards in a concentration-dependent manner. One trap sample tested positive for the presence of tetracyclines and sulphonamides. Taxonomic and resistome composition were largely maintained after storage with boric acid at -22 degrees C for up to five days.