Comprehensive Study of Antibiotics and Antibiotic Resistance Genes in Wastewater and Impacted Mediterranean Water Environments

Background: The spread of antimicrobial resistance is a central public health problem. Wastewater treatment plants and impacted environments are well-known hotspots for antibiotic resistance. However, there is still limited knowledge regarding where antibiotic resistance genes (ARGs) acquire mobility. Method: In this study, we aimed to gather evidence on the seasonal patterns of ARG spread in two Mediterranean areas from NE and E of Spain (Ebro River and Ebro Delta, and Xúquer River and Albufera de València), correlating ARG presence, with special focus on the faecal bacteria Escherichia coli, with antibiotic residues and environmental conditions. The analytical methodology employed was based on a suspect screening approach, while a novel prioritisation approach for antibiotics was proposed to identify those areas more susceptible to the spread of ARG. Results: Our findings demonstrate that ARG levels in wastewater were similar across different seasons, although a greater diversity of ARGs was recorded in summer. We hypothesise that horizontal gene transfer among aquatic bacterial populations during the northeastern Mediterranean summer, when temperatures reach approximately 35~40 °C, could be a key driver of ARG dissemination. By contrast, the highest concentrations of antibiotics in winter samples, with temperatures around 5~10 °C, may promote the spread of microbial resistance. Conclusions: Our key findings highlight that water temperature and sunlight irradiation are crucial factors influencing antibiotic levels and microbial abundance, requiring further investigation in future studies.