Fitness and Recovery of Bacterial Communities and Antibiotic Resistance Genes in Urban Wastewaters Exposed to Classical Disinfection Treatments

Abstract

Antibiotic resistance genes (ARGs) are increasingly appreciated to be important as micropollutants. Indirectly produced by human activities, they are released into the environment, as they are untargeted by conventional wastewater treatments. In order to understand the fate of ARGs and of other resistant forms (e.g., phenotypical adaptations) in urban wastewater treatment plants (WWTPs), we monitored three WWTPs with different disinfection processes (chlorine, peracetic acid (PAA), and ultraviolet light (UV)). We monitored WWTPs influx and pre- and postdisinfection effluent over 24 h, followed by incubation experiments lasting for 96 h. We measured bacterial abundance, size distribution and aggregational behavior, the proportion of intact (active) cells, and the abundances of four ARGs and of the mobile element integron1. While all the predisinfection treatments of all WWTPs removed the majority of bacteria and of associated ARGs, of the disinfection processes only PAA efficiently removed bacterial cells. However, the stress imposed by PAA selected for bacterial aggregates and, similarly to chlorine, stimulated the selection of ARGs during the incubation experiment. This suggests disinfections based on chemically aggressive destruction of bacterial cell structures can promote a residual microbial community that is more resistant to antibiotics and, given the altered aggregational behavior, to competitive stress in nature.

Peracetic acid disinfection kinetics for combined sewer overflows: indicator organisms, antibiotic resistance genes, and microbial community

“…Live-dead staining with flow cytometry was previously used to demonstrate the membrane destroying mechanism of PAA for WW disinfection. 15 At the beginning of treatment in the current study, there were no differences between total and viable ARG concentrations indicating ARGs were predominantly in viable cells, consistent with reports that bacterial cells with intact membranes comprised approximately 70% of the bacterial community in municipal wastewater influent. 15 A previous study differentiated intra- and extracellular ARGs in environmental matrices using a washing technique, observing extracellular DNA generally made up only less than 0.5% of total DNA in animal sludge samples.…”

“…15 At the beginning of treatment in the current study, there were no differences between total and viable ARG concentrations indicating ARGs were predominantly in viable cells, consistent with reports that bacterial cells with intact membranes comprised approximately 70% of the bacterial community in municipal wastewater influent. 15 A previous study differentiated intra- and extracellular ARGs in environmental matrices using a washing technique, observing extracellular DNA generally made up only less than 0.5% of total DNA in animal sludge samples. 40 In contrast, extracellular ARGs were found in higher concentration than intracellular ARGs in river sediments including those receiving treated wastewater.…”

“…This result is consistent with reports that PAA did not enhance overall ARG removal (i.e., reduce total ARG concentrations measured by qPCR) in post-secondary treated WW effluent after disinfection. 15,16 The application of vPCR in this study incorporates the specificity of qPCR and overcomes the limitation of qPCR to differentiate between DNA originating from viable and nonviable sources. PAA’s mode of action is presumed to be oxidization of sulfhydryl and sulfur bonds in proteins, enzymes and other metabolites, including those in microbial membranes.…”

“…Two recent studies demonstrated that PAA is not effective at reducing the total concentrations of tet (A), bla TEM , qnr S, erm B, sul 1, and sul 2 genes encoding resistance to tetracycline, beta-lactams, quinolone, erythromycin and sulfonamides respectively, in secondary WW effluents. 15, 16 Flow cytometry analysis with live-dead staining demonstrated damage to cell walls indicating that a portion of detected ARGs were extracellular or present in nonviable cells. 15 Little data is available reporting disinfection kinetics for ARG carrying cells in WW, 17 and none is available for CSO effluent.…”

“…15, 16 Flow cytometry analysis with live-dead staining demonstrated damage to cell walls indicating that a portion of detected ARGs were extracellular or present in nonviable cells. 15 Little data is available reporting disinfection kinetics for ARG carrying cells in WW, 17 and none is available for CSO effluent. Understanding the partitioning of ARGs between viable and non-viable sources is critical for understanding the mechanisms and therefore the risk of ARG proliferation in the environment.…”