An integrated One Health framework for holistic evaluation of risks from antifungal agents in a large-scale multi-city study using post-acquisition mass spectral data mining and wastewater-based epidemiology pipelines
Authors: Wattanayon Rawiwan , Proctor Kathryn , Jagadeesan Kishore Kumar , Barden Ruth , Kasprzyk-Hordern Barbara 
Published: March 2023

A new analytical framework for retrospective mass spectral data mining for antifungal (AFs) agents was developed as part of One Health framework for holistic evaluation of risks from AFs. Post-acquisition data mining of mass spectral repository obtained with a maXis HD QToF mass spectrometer, retrospective quantification and confirmation using bbCID method was undertaken. A large scale, multi-city spatiotemporal profiling programme of antifungal agents was undertaken in the Avon River catchment, South-West England encompassing five contrasting towns/cities served by five major wastewater treatment plants (WWTPs) covering a large geographical area of 2000 km2 and a population of ~1.5 million accounting for > 75% of the overall population in the studied catchment. Key drivers of AFs in the catchment were identified with communal WWTP discharges being the main driver for human AFs (e.g., fluconazole and ketoconazole) and agricultural runoff being the main driver for pesticide AFs (e.g., prochloraz, prothioconazole and tebuconazole). Daily mass loads (DLs) of AF were study/city dependent with several instances of large quantities of AFs (direct disposal) observed. Wastewater-Based Epidemiology (WBE) pipelines were developed to estimate community-wide exposure to antifungal agents and to triangulate the WBE data with per-postcode prescription. Average WBE-estimated human used fluconazole and ketoconazole PNDIs (population normalised daily intake per person) exceeded on average 300 mg day-1 1000inh-1 and 2000 mg day-1 1000inh-1. This is much higher than PNDPs (population normalised daily prescriptions) <40 mg day-1 1000inh-1 and <80 mg day-1 1000inh-1 for fluconazole and ketoconazole respectively. This was expected due to both prescription and OTC (over-the-counter) usage as well as both oral and topical (non-metabolic) application. WBE applied to pesticide-based AFs: prothioconazole and tebuconazole provided variable PNDI estimates across the catchment. Prothioconazole PNDIs were <40,000 mg day-1 1000inh -1, which gave intake per kg of body weight at: 0.43, 0.26, 0.07 mg kg-1 in City A, B and C and is likely due to accounting for external/non-human sources. This is higher than the acceptable daily intake (ADI) set at 0.01 mg kg-1bw day-1, which warrants further study. Intake per kg of body weight estimated using tebuconazole as a xenobiotic chemical residue (XCR) was: 0.86, 1.39, 0.12, 0.13 and 2.7 mg kg-1 in City A-E respectively and is likely due to accounting for external/non-human sources. Intake calculated using the metabolite is in line with ADI: 0.02 and 0.01 mg kg-1 in City B and C respectively, which aligns well with ADI set at 0.03 mg kg-1bw day-1. The environmental risk assessment of the target AAs indicates low/medium risk from fluconazole, prochloraz and tebuconazole, medium risk from epoxiconazole, prothioconazoles metabolite, and tebuconazole, and high risk for prothioconazole in river water. High risk was estimated from fluconazole, epoxiconazole, prothioconazole and its metabolite, tebuconazole, ketoconazole in influent (and/or effluent) samples, which is important due to possible raw sewage discharge via sewer overflows. Due to lack of information on PNEC (predicted no-effect concentration) for other AFs, risk assessment could not be undertaken, which warrants further study.