Impact of Redox Conditions on Antibiotic Resistance Conjugative Gene Transfer Frequency and Plasmid Fate in Wastewater Ecosystems

Abstract

Wastewater is a common pathway for the spread of antibiotic resistance (AR) genes and bacteria into the environment. Biological treatment can mitigate this path, but horizontal gene transfer (HGT) between bacteria also occurs in such processes, although the influence of bioreactor habitat and ecology on HGT frequency is not well understood. Here, we quantified how oxidation-reduction (redox) conditions impact the fate of a Green fluorescent protein (Gfp)-tagged AR plasmid (pRP4-gfp) within an E. coli host (EcoFJ1) in the liquid and biofilms of multi-phase bioreactors. Replicate bioreactors treating domestic wastewater were operated under stable aerobic (+195 ± 25 mV), anoxic (-15 ± 50 mV), and anaerobic (-195 ± 15 mV) conditions, and flow cytometry and selective plating were used to quantify donor strain, EcoFJ1(pRP4-gfp), and putative transconjugants over time. Plasmid pRP4-gfp-bearing cells disappeared rapidly in aerobic ecosystems (~2.0 log reduction after 72 h), especially in the liquid phase. Conversely, EcoFJ1(pRP4-gfp) and putative transconjugants persisted longest in anaerobic biofilms (~1.0 log reduction, after 72 h). Apparent plasmid transfer frequencies also were higher under anaerobic conditions. Further, protozoan abundances were over 20 times higher in aerobic reactors relative to anaerobic reactors, and protozoa numbers significantly inversely correlated with pRP4-gfp signals across all reactors (p < 0.05). Overall, HGT frequency was consequentially impacted by bioreactor habitat conditions and trophic effects, especially relative oxygen level and evident predation. These observations provide guidance for new and alternate bioreactor designs aimed at minimizing AR HGT during biological wastewater treatment.

Extended-Spectrum β-Lactamase and Carbapenemase Genes are Substantially and Sequentially Reduced during Conveyance and Treatment of Urban Sewage

“…As sewage travels through the UWS, the microbial communities and their ARGs are diluted, mixed with resident communities, 15,16 exposed to varying environmental conditions, and sequentially subject to both stochastic (e.g., dispersal and drift) and deterministic selective processes (e.g., by differing nutrient and redox conditions). 17,18 These processes can lead to the loss of some ARGs if their hosts are driven to extinction, whereas transfer of mobile ARGs can increase their likelihood of persistence, even though the original intestinal host bacteria may die off. It is therefore essential to track ARGs together with the community structure and mobility potential to help identify the processes that mitigate or favor antibiotic resistance across the entirety of the UWS.…”

“…Realizing that UWSs consist of compartments with very different physical–chemical conditions, efforts to describe ARG dynamics have mainly considered one or a few of these compartments at a time, such as the influent, the biological treatment reactor, and the effluent. ,,, However, these efforts fail to capture a full picture of UWSs, and rigorous studies on the occurrence of ARGs across the different compartments are absent, hampering an integrative understanding of the fate of ARGs from raw sewage, through conveyance and treatment processes, until final discharge. As sewage travels through the UWS, the microbial communities and their ARGs are diluted, mixed with resident communities, , exposed to varying environmental conditions, and sequentially subject to both stochastic (e.g., dispersal and drift) and deterministic selective processes (e.g., by differing nutrient and redox conditions). , These processes can lead to the loss of some ARGs if their hosts are driven to extinction, whereas transfer of mobile ARGs can increase their likelihood of persistence, even though the original intestinal host bacteria may die off. It is therefore essential to track ARGs together with the community structure and mobility potential to help identify the processes that mitigate or favor antibiotic resistance across the entirety of the UWS.…”

Tailoring CRISPR-Cas Immunity for the Degradation of Antibiotic Resistance Genes

“…However, detection of various types of ARGs has been reported in and near wastewater treatment facilities, [4][5][6][7] indicating them potential sources for ARGs at the same time. To address this rising challenge, the fate of ARGs in engineered and natural systems has been interrogated, [8][9][10][11][12][13] and remediation methods have been developed. [14][15][16] Most of the current methods remove ARGs as biomolecules based on the physiochemistry of DNAs.…”

“…It, unfortunately, seems inevitable as the ARGs may be required and accumulate for the removal of antibiotics in the wastewater from pharmaceutical industries, hospitals, or even municipal sewerage systems, leading to a dilemma in addressing ARG problems. To tackle this challenge, environmental scientists have interrogated the fate of ARGs in engineered and natural systems (9)(10)(11)(12), and have been developing remediation methods (13)(14)(15). For current environmental applications, most of the methods remove ARGs as biomolecules based on the physiochemistry of DNAs (13).…”

Extended spectrum β-lactamase and carbapenemase genes are substantially and sequentially reduced during conveyance and treatment of urban sewage

“…However, these efforts fail to capture a full picture of UWS and rigorous studies on the occurrence of ARGs across the different compartments are absent, hampering an integrative understanding of the fate of ARGs from raw sewage, through conveyance, treatment processes, and until final discharge. As sewage travels through the UWS, the microbial communities and their ARGs are diluted, mixed with resident communities 15,16 , exposed to varying environmental conditions and, being sequentially subject to both stochastic (e.g., dispersal and drift) and deterministic selective processes (e.g., by differing nutrient and redox conditions) 17,18 . These processes can lead to the loss of some ARGs if their hosts are driven to extinction, whereas transfer of mobile ARGs can increase their likelihood of persistence, even though the original intestinal host bacteria may die off.…”