Urban streams in various parts of the world are facing increased anthropogenic pressure on their water quality, and storm flow events represent one such source of complex physical, chemical, and biological perturbations. Microorganisms are important components of these streams from both ecological and public health perspectives. Analysis of the effect of perturbations on the stream microbial community can help improve current knowledge on the impact such chronic disturbances can have on these water resources. This study examines microbial community dynamics during rain-induced storm flow conditions in an urban stream of the Chicago Area Waterway System. Additionally, using shotgun metagenomics we identified significant shifts in the microbial community composition and functional gene content following a high-rainfall event, with potential environment and public health implications. Previous work in this area has focused on specific genes/organisms or has not assessed immediate storm flow impact.

Background In the United States, an estimated 14,748 wastewater treatment plants (WWTPs) provide wastewater collection, treatment, and disposal service to more than 230 million people. The quality of treated wastewater is often assessed by the presence or absence of fecal indicator bacteria. UV disinfection of wastewater is a common final treatment step used by many wastewater treatment plants in order to reduce fecal coliform bacteria and other pathogens; however, its potential impacts on the total effluent bacterial community are seemingly varied. This is especially important given that urban WWTPs typically return treated effluent to coastal and riverine environments and thus are a major source of microorganisms, genes, and chemical compounds to these systems. Following rainfall, stormflow conditions can result in substantial increases to effluent flow into combined systems. Methods Here, we conducted a lab-scale UV disinfection on WWTP effluent using UV dosage of 100 mJ/cm2 and monitored the active microbiome in UV-treated effluent and untreated effluent over the course of 48 h post-exposure using 16S rRNA sequencing. In addition, we simulated stormflow conditions with effluent UV-treated and untreated effluent additions to river water and compared the microbial communities to those in baseflow river water. We also tracked the functional profiles of genes involved in tetracycline resistance (tetW) and nitrification (amoA) in these microcosms using RT-qPCR. Results We showed that while some organisms, such as members of the Bacteroidetes, are inhibited by UV disinfection and overall diversity of the microbial community decreases following treatment, many organisms not only survive, but remain active. These include common WWTP-derived organisms such as Comamonadaceae and Pseudomonas. When combined with river water to mimic stormflow conditions, these organisms can persist in the environment and potentially enhance microbial functions such as nitrification and antibiotic resistance.

22Urban streams are susceptible to stormwater and sewage inputs that can impact their ecological 23 health and water quality. Microbial communities in streams play important functional roles and 24 their composition and metabolic potential can help assess ecological state and water quality. 25Although these environments are highly heterogenous, little is known about the influence of 26 isolated perturbations, such as those resulting from rain events on urban stream microbiota. Here, 27we examined the microbial community composition and diversity in an urban stream during dry 28 and wet weather conditions with both 16S rRNA gene sequencing across multiple years and 29shotgun metagenomics to more deeply analyze a single stormflow event. Metagenomics was 30 used to assess population-level dynamics as well as shifts in the microbial community taxonomic 31 profile and functional potential before and after a substantial rainfall. Results demonstrated 32 general trends present in the stream under stormflow vs. baseflow conditions across years and 33 seasons and also highlighted the significant influence of increased effluent flow following rain in 34 shifting the stream microbial community from abundant freshwater taxa to those more associated 35 with urban/anthropogenic settings. Shifts in the taxonomic composition were also linked to 36 changes in functional gene content, particularly for transmembrane transport and organic 37 substance biosynthesis. We also observed an increase in relative abundance of genes encoding 38 degradation of organic pollutants and antibiotic resistance after rain. Overall, this study provided 39 evidence of stormflow impacts on an urban stream microbiome from an environmental and 40 public health perspective. 41

Background. In the United States, an estimated 14,748 wastewater treatment plants (WWTPs) provide wastewater collection, treatment, and disposal service to more than 230 million people. The quality of treated wastewater is often assessed by the presence or absence of fecal indicator bacteria. UV disinfection of wastewater is a common final treatment step used by many wastewater treatment plants in order to reduce fecal coliform bacteria and other pathogens; however, its potential impacts on the total effluent bacterial community are seemingly varied. This is especially important given that urban wastewater treatment plants (WWTPs) typically return treated effluent to coastal and riverine environments and thus are a major source of microorganisms, genes, and chemical compounds to these systems. Following rainfall, stormflow conditions can result in substantial increases to effluent flow into these systems. Methods. Here, we conducted a lab-scale UV disinfection on WWTP effluent using UV dosage of 100 mJ/cm2 and monitored the active microbiome in UV-treated effluent and untreated effluent over the course of 48h post-exposure using 16S rRNA sequencing. In addition, we simulated stormflow conditions with effluent UV-treated and untreated effluent additions to river water and compared the microbial communities to those in baseflow river water. We also tracked the functional profiles of genes involved in tetracycline resistance (tetW) and nitrification (amoA) in these microcosms using qPCR. Results. We showed that while some organisms, such as members of the Bacteroidetes, are inhibited by UV disinfection and overall diversity of the microbial community decreases following treatment, many organisms not only survive, but remain active. These include common WWTP-derived organisms such as Comamonadaceae and Pseudomonas. When combined with river water to mimic stormflow conditions, these organisms can persist in the environment and potentially enhance microbial functions such as nitrification and antibiotic resistance.

Background. In the United States, an estimated 14,748 wastewater treatment plants (WWTPs) provide wastewater collection, treatment, and disposal service to more than 230 million people. The quality of treated wastewater is often assessed by the presence or absence of fecal indicator bacteria. UV disinfection of wastewater is a common final treatment step used by many wastewater treatment plants in order to reduce fecal coliform bacteria and other pathogens; however, its potential impacts on the total effluent bacterial community are seemingly varied. This is especially important given that urban wastewater treatment plants (WWTPs) typically return treated effluent to coastal and riverine environments and thus are a major source of microorganisms, genes, and chemical compounds to these systems. Following rainfall, stormflow conditions can result in substantial increases to effluent flow into these systems. Methods. Here, we conducted a lab-scale UV disinfection on WWTP effluent using UV dosage of 100 mJ/cm2 and monitored the active microbiome in UV-treated effluent and untreated effluent over the course of 48h post-exposure using 16S rRNA sequencing. In addition, we simulated stormflow conditions with effluent UV-treated and untreated effluent additions to river water and compared the microbial communities to those in baseflow river water. We also tracked the functional profiles of genes involved in tetracycline resistance (tetW) and nitrification (amoA) in these microcosms using qPCR. Results. We showed that while some organisms, such as members of the Bacteroidetes, are inhibited by UV disinfection and overall diversity of the microbial community decreases following treatment, many organisms not only survive, but remain active. These include common WWTP-derived organisms such as Comamonadaceae and Pseudomonas. When combined with river water to mimic stormflow conditions, these organisms can persist in the environment and potentially enhance microbial functions such as nitrification and antibiotic resistance.