Monitoring of antibiotics, pollutants, and AMR in formal and informal settlements: Back-calculation of antibiotic usage in the absence of prescription data

Antimicrobial resistance (AMR) is a major global topic, concerning the increasing pathogenic tolerance to antibiotics. Excessive and inappropriate antibiotic use heightens the emergence of antimicrobial resistance genes in pathogenic organisms, resulting in reduced drug susceptibility. The usage, excretion, and fate of antibiotics is an important aspect of AMR dissemination. In regions where prescription data / pharmacy records are not collated or publically available, an estimation of usage via back calculation from environmental concentrations can be vital. A novel analytical method was developed for broad- and narrowspectrum antibiotics, via UPLC-ESI-TQD-MS/MS. Longitudinal monitoring of these 59 drugs and 25 drug metabolites, other pollutants, and metagenomic data, is used to aid our understanding of AMR dissemination. River and wastewater samples (both aqueous and solid) were collected from 10 sites within Stellenbosch town, SA, across 7 sampling campaigns between May 2018 and June 2019. Estimation of antibiotic usage requires several parameters, including flow data to establish daily loads; knowledge of prescribed drug compositions; percentage drug elimination; route and mechanism of elimination; and the drug’s physio-chemical fate (e.g. solid/liquid partitioning). These estimations are achieved using antibiotic concentrations from >500 aqueous samples and >60 solid samples, together with the analysis of spatial and temporal trends. However, environmental drug persistence and the presence of other pollutants, such as heavy metals and biocides, are equally important to monitor – particularly when considering antimicrobial resistance. Exposure to sub-inhibitory concentrations of antibiotics is likely to impose selection favouring the presence of AMR genes; and acquisition through lateral gene transfer can lead to the emergence of AMR genes in the environment via co-selection. Subsequently, incidences of AMR dissemination are established by correlating the antibiotic and metagenomic data. Regulated and appropriate antibiotic usage is vital for controlling the spread of AMR. In the absence of records, wastewater based epidemiology is used to back calculate these data from the main town (via wastewater influent) as well as informal settlements without waste infrastructure (via pseudo-wastewater river samples). Acknowledgements We would like to acknowledge the Wolfaardt group, (Stellenbosch University) for sample collection, processing, and shipment to the University of Bath; the Stellenbosch University Central Analytical Facilities team (ICP-MS & XRF unit) for assistance with the ICP sample preparation and instrumental analysis; the members of the ReNEW project research team (www.gcrf-renew.co.uk); and our funders GCRF (Global Challenges Research Fund).