Degradation and horizontal gene transfer analysis of plasmid-encoded antibiotic resistance genes during UV254, hydroxyl radical and sulphate radical treatments

Abstract

The environment has been implicated with antibiotic resistance (AR). Effective degradation and deactivation of antibiotic resistance genes (ARGs) in wastewater treatment can serve as barriers to AR dissemination. This study investigated the degradation kinetics of intracellular (i-) and extracellular (e-) plasmid-encoded tetA, ampR and sul1 ARGs using UV254, hydroxyl radical (HO.) and sulphate radical (SO4.-) UV-based advanced oxidation processes (UV254/H2O2 and UV254/S2O82- respectively). The degradation of tetA, ampR and sul1 was quantified using quantitative polymerase chain reaction (qPCR). The damages to each ARG were observed using two qPCR amplicons ranging between 162 and 1054 bp. Culture-based horizontal gene transformation experiments were used to estimate the deactivation kinetics of pCR™2.1-TOPO AR plasmid. ARG degradation and deactivation kinetics were then compared to understand the roles of each treatment strategy in AR mitigation. Results indicate that extracellular ARGs degradation kinetics of the treatments followed an order UV254/S2O82- > UV254/H2O2 > UV254. The base pair specific kinetic constants with respect to HO. and SO4.- at pH 7 were between 1.86 × 109-1.65 × 1011 M−1s−1 and 2.87 × 109-5.84 × 1011 M−1s−1 respectively. e-ARGs degradation was at least 2-fold higher than i-ARGs degradation for all treatments. UV254/S2O82- was most effective for ARG degradation under acidic pH (5–6) while UV254/H2O2 was most effective between pH 7 and 8. Higher degradation rates were recorded for AT-rich ampR and longer qPCR amplicons. Deactivation rates by UV254/H2O2 and UV254/S2O82- were 2.6-times higher than that of UV254. Generally, deactivation kinetics were ∼ 8–13 times faster than degradation kinetics observed for short ampR amplicon. These findings show an overestimation of the potential risks of ARG presence using short qPCR target amplicons and the impact of nucleotide composition on ARG damage.