Microbe-sediment interactions in Great Lakes recreational waters: Implications for human health risk.

Microbial assessments of recreational water have traditionally focused on culturing or DNA-based approaches of the planktonic water column, omitting influence from microbe-sediment relationships. Sediment (bed and suspended) has been shown to often harbour levels of bacteria higher than the planktonic phase. The fate of suspended sediment (SS) bacteria is extensively related to transport dynamics (e.g., deposition) of the associated sediment/floc. When hydraulic energy allows, SS will settle, introducing new (potentially pathogenic) organisms to the bed. With turbulence, including waves, currents and swimmers, the risk of human ingestion is elevated due to resuspension of bed sediment and associated microbes. This research used multiplex nanofluidic reverse transcriptase quantitative PCR on RNA of bacteria associated with bed and SS to explore the active bacteria in freshwater shorelines. Bacterial genes of human health concern regarding recreational water use were targeted, such as faecal indicator bacteria (FIB), microbial source tracking genes and virulence factors from waterborne pathogens. Results indicate avian sources (i.e., gulls, geese) to be the largest nonpoint source of FIB associated with sediment in Great Lakes shorelines. This research introduces a novel approach to microbial water quality assessments and enhances our understanding of microbe-sediment dynamics and the quality of freshwater beaches.

Biogeochemical Processes and Microbial Dynamics Governing Phosphorus Retention and Release in Sediments: A Case Study in Lower Great Lakes Headwaters.

The ability of headwater bed and suspended sediments to mitigate non-point agricultural phosphorus (P) loads to the lower Great Lakes is recognized, but the specific biogeochemical processes promoting sediment P retention or internal P release remain poorly understood. To elucidate these mechanisms, three headwater segments located within priority watersheds of Southern Ontario, Canada, were sampled through the growing season of 2018-2020. The study employed equilibrium P assays along with novel assessments of legacy watershed nutrients, nitrogen (N) concentrations, sediment redox, and microbial community composition. 20-year data revealed elevated total P (TP) and total Nitrogen (TN) at an inorganic fertilizer and manure fertilizer-impacted site, respectively. Overall, sampled sites acted as P sinks; however, agricultural sediments exhibited significantly lower buffering capacity compared to a reference forested watershed. Collection of fine suspended sediment (<63 µm) through time-integrated sampling showed the suspended load at the inorganic-fertilized site was saturated with P, indicating a greater potential for P release into surface waters compared to bed sediments. Through vertical microsensor profiling and DNA sequencing of the sediment microbial community, site-specific factors associated with a distinct P-source event were identified. These included rapid depletion of dissolved oxygen (DO) across the sediment water interface (SWI), as well as the presence of nitrate-reducing bacterial and ammonia-oxidizing archaeal (AOA) genera. This research provides valuable insights into the dynamics of P in headwaters, shedding light on P retention and release. Understanding these processes is crucial for effective management strategies aimed at mitigating P pollution to the lower Great Lakes.

Impact of sample collection on prokaryotic and eukaryotic diversity of niche environments of the oil-sand mining impacted Athabasca River.

Microbial communities are an important aspect of overall riverine ecology; however, appreciation of the effects of anthropogenic activities on unique riverine microbial niches, and how the collection of these samples affects the observed diversity and community profile is lacking. We analyzed prokaryotic and eukaryotic communities from surface water, biofilms, and suspended load niches along a gradient of oil sands-related contamination in the Athabasca River (Alberta, Canada), with suspended load or particle-associated communities collected either via Kenney Sampler or centrifugation manifold. At the phylum level, different niche communities were highly similar to each other and across locations. However, there were significant differences in the abundance of specific genera among the different niches and across sampling locations. A generalized linear model revealed that use of the Kenney Sampler resulted in more diverse bacterial and eukaryotic suspended load community than centrifugal collection, though suspended load communities collected by any means remained stably diverse across locations. Although there was an influence of water quality parameters on community composition, all sampled sites support diverse bacterial and eukaryotic communities regardless of the degree of contamination, highlighting the need to look beyond ecological diversity as a means of assessing ecological perturbations, and consider collecting samples from multiple niche environments.

Exploring bacterial pathogen community dynamics in freshwater beach sediments: A tale of two lakes.

Pathogenic bacteria associated with freshwater ecosystems can pose significant health risks particularly where recreational water use is popular. Common water quality assessments involve quantifying indicator Escherichia coli within the water column but neglect to consider physical and geochemical factors and contributions from the sediment. In this study, we used high-throughput sequencing to investigate sediment microbial communities at four freshwater public beaches in southern Ontario, Canada and analysed community structure, function, and gene expression with relation to geographical characteristics. Our results indicate that beach sediments at the sediment-water interface could serve as potential sources of bacterial contamination under low-energy environments with tightly packed small sediment particles compared with high-energy environments. Further, the absence of pathogens but expression of pathogenic transcripts suggests occurrence of alternate gene acquisition. Pathogenicity at these locations included expression of Salmonella virulence factors, genes involved in pertussis, and antimicrobial resistance. Finally, we introduce a proposed universal bacterial pathogen model to consider the combined and synergistic processes used by these microbes. To our knowledge, this is the first study of its kind to investigate chemolithotrophic activity related to pathogens within bed sediment at freshwater beaches. This work helps advance current understanding of health risks in these environments.

Influence of wave action on the partitioning and transport of unattached and floc-associated bacteria in fresh water.

The dynamic interaction of bacteria within bed sediment and suspended sediment (i.e., floc) in a wave-dominated beach environment was assessed using a laboratory wave flume. The influence of shear stress (wave energy) on bacterial concentrations and on the partitioning and transport of unattached and floc-associated bacteria was investigated. The study showed that increasing wave energy (0.60 and 5.35 N/s) resulted in a 0.5 to 1.5 log increase in unattached cells of the test bacterium Pseudomonas sp. strain CTO7::gfp-2 in the water column. There was a positive correlation between the bacterial concentrations in water and the total suspended solids, with the latter increasing from values of near 0 to up to 200 mg/L over the same wave energy increase. The median equivalent spherical diameter of flocs in suspension also increased by an order of magnitude in all experimental trials. Under both low (0.60 N/s) and high (5.35 N/s) energy regime, bacteria were shown to preferentially associate with flocs upon cessation of wave activity. The results suggest that collecting water samples during periods of low wave action for the purpose of monitoring the microbiological quality of water may underestimate bacterial concentrations partly because of an inability to account for the effect of shear stress on the erosion and mobilization of bacteria from bed sediment to the water column. This highlights the need to develop a more comprehensive beach analysis strategy that not only addresses presently uncharacterized shores and sediments but also recognizes the importance of eroded flocs as a vector for the transport of bacteria in aquatic environments.

Diversity of integron- and culture-associated antibiotic resistance genes in freshwater floc.

Clinically important antibiotic resistance genes were detected in culturable bacteria and class 1 integron gene cassettes recovered from suspended floc, a significant aquatic repository for microorganisms and trace elements, across freshwater systems variably impacted by anthropogenic activities. Antibiotic resistance gene cassettes in floc total community DNA differed appreciably in number and type from genes detected in bacteria cultured from floc. The number of floc antibiotic resistance gene cassette types detected across sites was positively correlated with total (the sum of Ag, As, Cu, and Pb) trace element concentrations in aqueous solution and in a component of floc readily accessible to bacteria. In particular, concentrations of Cu and Pb in the floc component were positively correlated with floc resistance gene cassette diversity. Collectively, these results identify suspended floc as an important reservoir, distinct from bulk water and bed sediment, for antibiotic resistance in aquatic environments ranging from heavily impacted urban sites to remote areas of nature reserves and indicate that trace elements, particularly Cu and Pb, are geochemical markers of resistance diversity in this environmental reservoir. The increase in contamination of global water supplies suggests that aquatic environments will become an even more important reservoir of clinically important antibiotic resistance in the future.

Comparative floc-bed sediment trace element partitioning across variably contaminated aquatic ecosystems.

Significantly higher concentrations of Ag, As, Cu, Ni and Co are found in floc compared to bed sediments across six variably impacted aquatic ecosystems. In contrast to the observed element and site-specific bed sediment trace element (TE) partitioning patterns, floc TE sequestration is consistently dominated by amorphous oxyhydroxides (FeOOH), which account for 30-79% of floc total TE concentrations, irrespective of system physico-chemistry or elements involved. FeOOH consistently occur in significantly higher concentrations in floc than within bed sediments. Further, comparative concentration factors indicate significantly higher TE reactivity of floc-FeOOH relative to sediment-FeOOH in all systems investigated, indicating that both the greater abundance and higher reactivity of floc-FeOOH contribute to enhanced floc TE uptake. Results indicate that floc-organics (live cells and exopolymeric substances, EPS) directly predict floc-FeOOH concentrations, suggesting an organic structural role in the collection/templating of FeOOH. This, in turn, facilitates the sequestration of TEs associated with floc-FeOOH formation, imparting the conserved FeOOH "signature" on floc TE geochemistry across sites. Results demonstrate that the organic rich nature of floc exerts an important control over TE geochemistry in aquatic environments, ultimately creating a distinct solid with differing controls over TE behavior than bed sediments in close proximity (<0.5 m).

Identifying chemolithotrophic and pathogenic-related gene expression within suspended sediment flocs in freshwater environments: A metatranscriptomic assessment.

The introduction and proliferation of pathogenic organisms in aquatic systems is a serious global issue that consequently leads to economic, financial, and health concerns. Health and safety related to recreational water use is typically monitored through water quality assessments that are outdated and can be misleading. These traditional methods focus on broad taxa groups, provide no insight into the active community or source of contamination, and the sediment compartments (bed and suspended) are often overlooked. To bridge this knowledge gap, our study aimed to 1) examine the metatranscriptome of the microbial community associated with suspended sediment (SS) in freshwater systems; 2) explore the influence of SS in tributaries to the littoral zone of the receiving lake; and 3) compare the SS fraction with previously reported nearshore bed sediment data. Samples were collected seasonally from Lake St. Clair and Lake Erie. Beaches in this region are influenced by both agriculture runoff and continued urban expansion. Results show that both adjacent tributary and beach SS have similar microbial functional diversity and are strongly correlated by site and season. We identified expression of transcripts encoding sequences with similarities to genes involved in nine bacterial infectious disease pathways, including legionellosis (sdhA) and Vibrio cholerae pathogenesis. According to MG-RAST gene categories, lake samples typically showed higher overall expression (p < 0.05) of transcripts with similarities to genes involved in infectious disease pathways compared to the tributaries, with summer upregulated (p < 0.05) compared to fall. Our data suggests SS acts as a strong vector for pathogen transport, making this facet an important area for further research as it pertains to human health regarding recreational water use. To our knowledge, this work is the first to investigate SS in aquatic microbial communities using metatranscriptomic analyses and has significant potential to help address growing issues of microbial contamination impacting freshwater security.

Physical and ecological controls on freshwater floc trace metal dynamics.

Significantly higher concentrations of Ag, As, Cu, Co, Ni, and Pb are found in suspended floc compared to surficial bed sediments for a freshwater beach in Lake Ontario. Contrasting observed element-specific bed sediment metal partitioning patterns, floc sequestration for all elements is dominated by one substrate: amorphous oxyhydroxides. More specifically, floc metal scavenging is controlled by floc biogeochemical architecture. Floc organics, largely living microbial cells and associated exopolymeric substances (EPS), act as scaffolds for the collection and/or templating of amorphous Fe oxyhydroxides. While interactions between floc organics and amorphous Fe oxyhydroxides affected floc sorption behavior, specific element affinities and competition for these limited substrates was important for overall floc partitioning. Further, assessment of metal dynamics during stormy conditions indicated energy-regime driven shifts in floc and bed sediment partitioning that were specifically linked to the exchange of floc and bed sedimentary materials. These novel results demonstrate that the microbial nature of floc formation exerts an important control on floc metal dynamics distinguishable from surficial bed sediments and that hydrologic energy-regime is an important factor to consider in overall floc metal behavior, especially in beach environments.

A structure-function based approach to floc hierarchy and evidence for the non-fractal nature of natural sediment flocs.

Natural sediment flocs are fragile, highly irregular, loosely bound aggregates comprising minerogenic and organic material. They contribute a major component of suspended sediment load and are critical for the fate and flux of sediment, carbon and pollutants in aquatic environments. Understanding their behaviour is essential to the sustainable management of waterways, fisheries and marine industries. For several decades, modelling approaches have utilised fractal mathematics and observations of two dimensional (2D) floc size distributions to infer levels of aggregation and predict their behaviour. Whilst this is a computationally simple solution, it is highly unlikely to reflect the complexity of natural sediment flocs and current models predicting fine sediment hydrodynamics are not efficient. Here, we show how new observations of fragile floc structures in three dimensions (3D) demonstrate unequivocally that natural flocs are non-fractal. We propose that floc hierarchy is based on observations of 3D structure and function rather than 2D size distribution. In contrast to fractal theory, our data indicate that flocs possess characteristics of emergent systems including non-linearity and scale-dependent feedbacks. These concepts and new data to quantify floc structures offer the opportunity to explore new emergence-based floc frameworks which better represent natural floc behaviour and could advance our predictive capacity.

Effects of ultrasound on suspended particles in municipal wastewater.

The objective of this research is to explore the fundamental characteristics of how particles in wastewater respond to ultrasound, with an aim to improve wastewater disinfection. Particles of a predetermined size fraction and concentration were treated with varying doses of ultrasound at 20.3 kHz. Ultrasonic power transfer to the fluid was measured using calorimetry or acoustical measurements. Image analysis particle counting was used to measure the size distribution of particles before and after ultrasound treatment. The influence of three parameters: particle origin (raw wastewater or from the aeration basin of the activated sludge process), particle concentration, and particle size on the percentage of particle breakage after ultrasound treatment was compared. It was found that raw wastewater and aeration basin particles of the same size fraction (90-106 microm) responded to ultrasound in a similar way. Particle breakage was not affected by changes in particle concentration from 100 to 400 particles per mL. Larger wastewater particles (90-250 microm) were more susceptible to breakage than smaller ones (38-63 microm diameter). The percentage of particle breakage increased linearly with a logarithmic increase in the ultrasound energy density, that is the ultrasound energy delivered per unit volume of the sample (R(2)=0.48-0.91). An expression that predicts the percent of particles broken as a function of ultrasound energy density is provided.

The Alberta oil sands eroded bitumen/sediment transitional journey: Influence on sediment transport dynamics, PAH signatures and toxicological effect.

This paper investigated the sediment/contaminant continuum of bitumen containing sediment, from eroded exposed natural bitumen outcrops to river depositional zone, in order to improve our understanding of the transitional sediment, chemistry and toxicological influence on aquatic health. To achieve this aim, we linked a rainfall simulator with an annular flume to allow for connectivity between terrestrial erosion to stream flow. Bulk sediments were collected from the minable McMurray Formation (MF) on the Ells (EL) and Steepbank (STB) Rivers and from the Clearwater Formation (CF) on the STB. All plots generated similar washoff rates (~25 L hr-1) and fine eroded sediment (d50 = 5 µm) regardless of strata. The CF generated the highest total eroded sediment (TES) followed by EL-MF and STB-MF. The STB-MF generated up to three orders of magnitude lower TES than the STB-CF. The EL generated the highest PAH concentrations, however, when normalizing by TES mass delivered to the flume, STB-MF PAH was one to two orders of magnitude higher per unit mass than EL-MF. The TES concentrations were inversely proportional to the degree of bitumen within the sediment (STB-MF > EL-MF > STB-CF). Once the TES was suspended in the flume, there was a change in the dominant PAH for most classes [e.g., Dibenzothiophenes C4 (parent material) to a C3 (TES)]. Further, no flocculation and the buoyant properties of the TES suggested that the sediment and associated PAHs may travel long distances. The EL-MF proved to be the most toxic to fathead minnow embryo survival due principally to the high PAH concentrations and sediment loads. The CF exhibited no toxicological effect. This work has shown the importance of assessing sediment/contaminant characteristics over the continuum (terrestrial erosion/washoff to river deposition) in order to support basin wide management strategies for the protection of aquatic health.

Microbial metabolic strategies for overcoming low-oxygen in naturalized freshwater reservoirs surrounding the Athabasca Oil Sands: A proxy for End-Pit Lakes?

The success and sustainability of aquatic ecosystems are driven by the complex, cooperative metabolism of microbes. Ecological engineering strategies often strive to harness this syntrophic synergy of microbial metabolism for the reclamation of contaminated environments worldwide. Currently, there is a significant knowledge gap in our understanding of how the natural microbial ecology overcomes thermodynamic limitations in recovering contaminated environments. Here, we used in-situ metatranscriptomics and associated metataxonomic analyses on sediments collected from naturalized freshwater man-made reservoirs within the Athabasca Oil Sands region of Alberta, Canada. These reservoirs are unique since they are untouched by industrial mining processes and serve as representative endpoints for model landscape reconstruction. Results indicate that a microbial syntrophic cooperation has been established represented by the oxygenic and anoxygenic phototrophs, sustained through the efficient use of novel cellular mechanistic adaptations tailored to these unique thermodynamic conditions. Specifically, chemotaxis transcripts (cheY & MCPs-methyl-accepting chemotaxis proteins) were highly expressed, suggesting a highly active microbial response to gradients in environmental stimuli, resulting indirectly from hydrocarbon compound alteration. A high expression of photosynthetic activity, likely sustaining nutrient delivery to the similarly highly expressed methanogenic community acting in syntrophy during the breakdown of organics. Overall the more diversified functionality within sub-oxic sample locations indicates an ability to maintain efficient metabolism under thermodynamic constraints. This is the first study to holistically identify and characterize these types of in-situ, metabolic processes and address their thermodynamic feasibility within a global context for large landscape reconstruction. These characterizations of regional, natural landscapes surrounding the oil sands mining operation are severely lacking, but truly provide invaluable insight into end-point goals and targets for reclamation procedures.

Modelling the potential effects of Oil-Sands tailings pond breach on the water and sediment quality of the Lower Athabasca River.

Within the Oil-Sands industry in Alberta, Canada, tailings ponds are used as water recycling and tailings storage facilities (TSF) for mining activities. However, there could be possible circumstances under which a sudden breach of an embankment confining one of the TSFs may occur. Such a tailings pond breach would result in a sudden release of a huge volume of Oil Sands process-affected water (OSPW) and sediment slurry containing substantial amount of chemical constituents that would follow the downstream drainage paths and subsequently enter into the Lower Athabasca River (LAR). This study investigates the implications of OS tailings release on the water and sediment quality of the LAR by simulating the fate of sediment and associated chemicals corresponding to a hypothetical breach and release scenarios from a select set of tailings ponds using a two-dimensional hydrodynamic and constituent transport model. After predicting the total volume, time evolution and concentration of sediment and associated chemicals (metals, polycyclic aromatic hydrocarbons (PAHs) and naphthenic acids (NAs)) reaching the LAR, the transport and deposition of these materials within the study reach is simulated. The results show that, depending on tailings release locations, between 40 and 70% of the sediment and associated chemicals get deposited onto the river bed of the 160 km study reach while the rest leaves the study domain during the first three days following the release event. These sediment/chemicals deposited during the initial spill may also have long-term effects on the water quality and aquatic ecosystem of the river and the downstream delta. However, care has to be taken in interpreting the results as further analysis has shown that the outcomes of such model simulations are very sensitive to the various underlying assumptions as well as the values assigned to some model parameters representing the physical properties of the tailings material.

Novel insights into freshwater hydrocarbon-rich sediments using metatranscriptomics: Opening the black box.

Baseline biogeochemical surveys of natural environments is an often overlooked field of environmental studies. Too often research begins once contamination has occurred, with a knowledge gap as to how the affected area behaved prior to outside (often anthropogenic) influences. These baseline characterizations can provide insight into proposed bioremediation strategies crucial in cleaning up chemical spill sites or heavily mined regions. Hence, this study was conducted to survey the in-situ microbial activity within freshwater hydrocarbon-rich environments cutting through the McMurray formation - the geologic strata constituting the oil sands. We are the first to report in-situ functional variations among these freshwater microbial ecosystems using metatranscriptomics, providing insight into the in-situ gene expression within these naturally hydrocarbon-rich sites. Key genes involved in energy metabolism (nitrogen, sulfur and methane) and hydrocarbon degradation, including transcripts relating to the observed expression of methane oxidation are reported. This information provides better linkages between hydrocarbon impacted environments, closing knowledge gaps for optimizing not only oil sands mine reclamation but also enhancing microbial reclamation strategies in various freshwater environments. These finding can also be applied to existing contaminated environments, in need of efficient reclamation efforts.

Temporal and spatial trends in riverine suspended sediment and associated polycyclic aromatic compounds (PAC) within the Athabasca oil sands region.

Bitumen-bearing suspended sediment (SS) eroded from the McMurray Formation (MF) are fine grained (silts and clays) and coated with natural hydrophobic oils. This results in poor settling and long range transport of associated contaminants. There was a longitudinal increase in polycyclic aromatic compound (PAC) concentrations for rivers that erode the MF from upstream to downstream regardless of time-of-year, while loads were substantially increased during high flow periods when the erosive forces are the greatest and the overland flow contribution is high. Within the MF, variation in PAC contributions is seen by the Ells River having higher loads than the Steepbank River. Using the Ells and Steepbank as examples, double plot PACs ratios suggest that the PAC concentrations and signatures vary spatially within the MF but that the weathering processes may be the same. Plots of the various homologs of PACs generally illustrated a normal distribution which suggests petrogenic origins. However, several PAC ratios suggested that both the parent material and the SS are pyrogenic in nature. While it is likely that some level of atmospheric deposition (anthropogenic or from forest fire) is incorporated into the SS of the rivers, it is likely to be limited relative to the proportion of naturally eroded MF sediments. Additional analysis will be needed to distinguish the relative risk of both anthropogenic (e.g., industrial operations) and natural sources (bitumen deposits, forest fire) of PACs to the SS and to long-range depositional environments, as they may have potential aquatic ecological effects.

Compartmentalization of metals within the diverse colloidal matrices comprising activated sludge microbial flocs.

Activated sludge floc from a wastewater treatment system was characterized, with regard to principal structural, chemical, and microbiological components and properties, in relation to contaminant-colloid associations and settling. Multiscale analytical microscopies, in conjunction with multimethod sample preparations, were used correlatively to characterize diverse colloidal matrices within microbial floc. Transmission electron microscopy, in conjunction with energy dispersive spectroscopy (EDS), revealed specific associations of contaminant heavy metals with individual bacterial cells and with extracellular polymeric substances (EPS). Floc structure was mapped from the level of gross morphology down to the nano-scale, and flocs were described with respect to settling properties, size, shape, density, porosity, bound water content, and EPS chemical composition; gross surface properties were also measured for correlation with principal floc features. Compartmentalization results based on 171 EDS analyses and representative high-resolution images showed that nano-scale agglomerations of (i) silver (100%) and (ii) zinc (91%) were confined almost entirely to EPS matrices while (iii) Pb (100%) was confined to intracellular granules and (iv) aluminum was partitioned between EPS matrices (41%) and intracellular matrices (59%). The results suggest that engineered changes in microbial physiology and/or in macromolecular EPS composition may influence metal removal efficiencies.

Dynamic existence of waterborne pathogens within river sediment compartments. Implications for water quality regulatory affairs.

The transport and fate of indicator E. coli and Salmonella are shown to be highly influenced by their relationship with flocculated suspended and bed sediment particles. Flocs were found to dominate the suspended sediment load and have the effect of increasing the downward flux of the sediment to the river bed. Bacteria counts were consistently higher within sediment compartments (suspended and bed) than for the water alone, with the bed sediment found to represent a possible reservoir of pathogens for subsequent remobilization and transport to potentially high risk areas. The mechanism of microbial attachment and entrapment within the sediment was strongly linked to the EPS fibrils secreted by the biological consortium of the aquatic system. It is suggested that the sediment/pathogen relationship should be of concern to public health officials because of its potential effects on pathogen source fate and effect with implications on public health risk assessment. Current standard sampling strategies, however, are based on an assumption that bacteria are entirely planktonic and do not account for the potentially significant concentration of bacteria from the sediment compartments. The lack of understanding around pathogen/sediment associations may lead to an inaccurate estimate of public health risk, and, as such, possible modification of sampling strategies to reflect this association may be warranted.