Unraveling tetracycline and its degradation product: Induction mechanisms of antibiotic resistance in Escherichia coli

In aquatic environments, antibiotics degrade into byproducts, potentially enhancing bacterial resistance. However, the specific mechanisms by which these byproducts induce bacterial resistance remain elusive. This study conducted experimental evolution experiments to explore how E. coli adapts to tetracycline (TC) and its primary degradation products—anhydrotetracycline (ATC), epitetracycline (ETC), and 4-epianhydrotetracycline (EATC)—through evolution experiments. Prolonged exposure to TC and its byproducts significantly increased frequency of resistant mutants in E. coli ATCC25922, with a maximum 10⁶-fold increase. Resistant mutants exhibited markedly elevated minimum inhibitory concentrations (MICs) for TC, ampicillin (AMP), and ciprofloxacin (CIP), indicating multidrug resistance. Transcriptomic analysis showed that the antibiotic resistance phenotype could be related to enhanced target protection, metabolic adaptations, and reduced membrane permeability. The induction pathways between TC and its byproducts were distinct. Specifically, TC_20d (where TC_20d represents the mutants collected after 20 days of continuous exposure to TC) was associated with more alterations in ribosome-associated genes, which was correlated with an enhanced defensive response as shown by the data. Moreover, variations in energy metabolism gene expression suggest a robust metabolic defense in ATC_20d and ETC_20d. When TC and its byproducts—ATC, ETC, and EATC—act together, they induce antibiotic resistant mutants at rates of 29.8 %, 18.9 %, 18.3 %, and 31.9 %, respectively. This study provides a descriptive overview of the possible adaptive mechanisms and pathways that may be involved in antibiotic resistance due to environmental exposure.