Table 2_Filtered wastewater effluent promotes antibiotic tolerance by affecting Escherichia coli growth dynamics under tetracycline stress.xlsx

Filtered wastewater effluent contains dissolved compounds that may alter microbial dynamics and promote antibiotic tolerance, enabling bacteria to survive transient antibiotic exposure without acquiring genetic resistance. We examined the physiological and transcriptomic responses of E. coli exposed to filtered wastewater effluent and tetracycline to characterize tolerance in relation to growth dynamics. Growth assays and RNA sequencing were performed following simultaneous exposure. No differences in minimum inhibitory concentration (MIC) values were observed between wastewater-exposed and control populations, indicating no acquisition of heritable resistance. However, wastewater-exposed Escherichia coli exhibited significantly shortened lag phases compared to nanopure water controls, demonstrating a growth advantage under tetracycline stress. Consistent with this accelerated exit from the lag phase of growth, transcriptomic analysis revealed fewer differentially expressed genes under combined wastewater effluent and tetracycline exposure, suggesting that cells required fewer gene expression changes to adapt to the stress compared to cells growing under tetracycline stress but in absence of filtered wastewater effluent. Together, these results indicate that dissolved constituents in filtered wastewater effluent enhanced bacterial growth and stress resilience during tetracycline exposure, consistent with antibiotic tolerance rather than resistance. This suggests that effluent chemistry plays a critical role in shaping bacterial persistence in aquatic environments impacted by both antibiotics and wastewater effluent, with important implications for wastewater management and ecosystem risk assessment.