Quantification of microcystin production and biodegradation rates in the western basin of Lake Erie.

Cyanobacterial biomass forecasts currently cannot predict the concentrations of microcystin, one of the most ubiquitous cyanotoxins that threaten human and wildlife health globally. Mechanistic insights into how microcystin production and biodegradation by heterotrophic bacteria change spatially and throughout the bloom season can aid in toxin concentration forecasts. We quantified microcystin production and biodegradation during two growth seasons in two western Lake Erie sites with different physicochemical properties commonly plagued by summer Microcystis blooms. Microcystin production rates were greater with elevated nutrients than under ambient conditions and were highest nearshore during the initial phases of the bloom, and production rates were lower in later bloom phases. We examined biodegradation rates of the most common and toxic microcystin by adding extracellular stable isotope-labeled microcystin-LR (1 µg L-1), which remained stable in the abiotic treatment (without bacteria) with minimal adsorption onto sediment, but strongly decreased in all unaltered biotic treatments, suggesting biodegradation. Greatest biodegradation rates (highest of -8.76 d-1, equivalent to the removal of 99.98% in 18 h) were observed during peak bloom conditions, while lower rates were observed with lower cyanobacteria biomass. Cell-specific nitrogen incorporation from microcystin-LR by nanoscale imaging mass spectrometry showed that a small percentage of the heterotrophic bacterial community actively degraded microcystin-LR. Microcystin production and biodegradation rates, combined with the microcystin incorporation by single cells, suggest that microcystin predictive models could be improved by incorporating toxin production and biodegradation rates, which are influenced by cyanobacterial bloom stage (early vs. late bloom), nutrient availability, and bacterial community composition.

Heterotrophic Bacteria Dominate Catalase Expression during Microcystis Blooms.

In the oligotrophic oceans, key autotrophs depend on "helper" bacteria to reduce oxidative stress from hydrogen peroxide (H2O2) in the extracellular environment. H2O2 is also a ubiquitous stressor in freshwaters, but the effects of H2O2 on autotrophs and their interactions with bacteria are less well understood in freshwaters. Naturally occurring H2O2 in freshwater systems is proposed to impact the proportion of microcystin-producing (toxic) and non-microcystin-producing (nontoxic) Microcystis in blooms, which influences toxin concentrations and human health impacts. However, how different strains of Microcystis respond to naturally occurring H2O2 concentrations and the microbes responsible for H2O2 decomposition in freshwater cyanobacterial blooms are unknown. To address these knowledge gaps, we used metagenomics and metatranscriptomics to track the presence and expression of genes for H2O2 decomposition by microbes during a cyanobacterial bloom in western Lake Erie in the summer of 2014. katG encodes the key enzyme for decomposing extracellular H2O2 but was absent in most Microcystis cells. katG transcript relative abundance was dominated by heterotrophic bacteria. In axenic Microcystis cultures, an H2O2 scavenger (pyruvate) significantly improved growth rates of one toxic strain while other toxic and nontoxic strains were unaffected. These results indicate that heterotrophic bacteria play a key role in H2O2 decomposition in Microcystis blooms and suggest that their activity may affect the fitness of some Microcystis strains and thus the strain composition of Microcystis blooms but not along a toxic versus nontoxic dichotomy. IMPORTANCE Cyanobacterial harmful algal blooms (CHABs) threaten freshwater ecosystems globally through the production of toxins. Toxin production by cyanobacterial species and strains during CHABs varies widely over time and space, but the ecological drivers of the succession of toxin-producing species remain unclear. Hydrogen peroxide (H2O2) is ubiquitous in natural waters, inhibits microbial growth, and may determine the relative proportions of Microcystis strains during blooms. However, the mechanisms and organismal interactions involved in H2O2 decomposition are unexplored in CHABs. This study shows that some strains of bloom-forming freshwater cyanobacteria benefit from detoxification of H2O2 by associated heterotrophic bacteria, which may impact bloom development.

Environmental factors affecting chytrid (Chytridiomycota) infection rates on Planktothrix agardhii.

Planktothrix agardhii dominates the cyanobacterial harmful algal bloom biomass in Sandusky Bay, Lake Erie (USA) from May until September. This filamentous cyanobacterium known parasites including the chytrid fungal species Rhizophydium sp. C02, which was previously isolated from this region. The purpose of our work has been to establish how parasitic interactions affect Planktothrix population dynamics during a bloom event. Samples analyzed from the 2015 to 2019 bloom seasons using quantitative PCR investigate the spatial and temporal prevalence of chytrid infections. Abiotic factors examined in lab include manipulating temperature (17-31°C), conductivity (0.226-1.225 mS/cm) and turbulence. Planktothrix-specific chytrids are present throughout the bloom period and are occasionally at high enough densities to exert parasitic pressure on their hosts. Temperatures above 27.1°C in lab can inhibit chytrid infection, indicating the presence of a possible upper thermal refuge for the host. Data suggest that chytrids can survive conductivity spikes in lab at levels three-fold above Sandusky Bay waters if given sufficient time (7-12 days), whereas increased turbulence in lab severely inhibits chytrid infections, perhaps due to disruption of chemical signaling. Overall, these data provide insights into the environmental conditions that inhibit chytrid infections during Planktothrix-dominated blooms in temperate waters.

The Lake Erie HABs Grab: A binational collaboration to characterize the western basin cyanobacterial harmful algal blooms at an unprecedented high-resolution spatial scale.

Monitoring of cyanobacterial bloom biomass in large lakes at high resolution is made possible by remote sensing. However, monitoring cyanobacterial toxins is only feasible with grab samples, which, with only sporadic sampling, results in uncertainties in the spatial distribution of toxins. To address this issue, we conducted two intensive "HABs Grabs" of microcystin (MC)-producing Microcystis blooms in the western basin of Lake Erie. These were one-day sampling events during August of 2018 and 2019 in which 100 and 172 grab samples were collected, respectively, within a six-hour window covering up to 2,270 km2 and analyzed using consistent methods to estimate the total mass of MC. The samples were analyzed for 57 parameters, including toxins, nutrients, chlorophyll, and genomics. There were an estimated 11,513 kg and 30,691 kg of MCs in the western basin during the 2018 and 2019 HABs Grabs, respectively. The bloom boundary poses substantial issues for spatial assessments because MC concentration varied by nearly two orders of magnitude over very short distances. The MC to chlorophyll ratio (MC:chl) varied by a factor up to 5.3 throughout the basin, which creates challenges for using MC:chl to predict MC concentrations. Many of the biomass metrics strongly correlated (r > 0.70) with each other except chlorophyll fluorescence and phycocyanin concentration. While MC and chlorophyll correlated well with total phosphorus and nitrogen concentrations, MC:chl correlated with dissolved inorganic nitrogen. More frequent MC data collection can overcome these issues, and models need to account for the MC:chl spatial heterogeneity when forecasting MCs.

Individual Microcystis colonies harbour distinct bacterial communities that differ by Microcystis oligotype and with time.

Interactions between bacteria and phytoplankton in the phycosphere have impacts at the scale of whole ecosystems, including the development of harmful algal blooms. The cyanobacterium Microcystis causes toxic blooms that threaten freshwater ecosystems and human health globally. Microcystis grows in colonies that harbour dense assemblages of other bacteria, yet the taxonomic composition of these phycosphere communities and the nature of their interactions with Microcystis are not well characterized. To identify the taxa and compositional variance within Microcystis phycosphere communities, we performed 16S rRNA V4 region amplicon sequencing on individual Microcystis colonies collected biweekly via high-throughput droplet encapsulation during a western Lake Erie cyanobacterial bloom. The Microcystis phycosphere communities were distinct from microbial communities in whole water and bulk phytoplankton seston in western Lake Erie but lacked 'core' taxa found across all colonies. However, dissimilarity in phycosphere community composition correlated with sampling date and the Microcystis 16S rRNA oligotype. Several taxa in the phycosphere were specific to and conserved with Microcystis of a single oligotype or sampling date. Together, this suggests that physiological differences between Microcystis strains, temporal changes in strain phenotypes, and the composition of seeding communities may impact community composition of the Microcystis phycosphere.

Roles of Nutrient Limitation on Western Lake Erie CyanoHAB Toxin Production.

Cyanobacterial harmful algal bloom (CyanoHAB) proliferation is a global problem impacting ecosystem and human health. Western Lake Erie (WLE) typically endures two highly toxic CyanoHABs during summer: a Microcystis spp. bloom in Maumee Bay that extends throughout the western basin, and a Planktothrix spp. bloom in Sandusky Bay. Recently, the USA and Canada agreed to a 40% phosphorus (P) load reduction to lessen the severity of the WLE blooms. To investigate phosphorus and nitrogen (N) limitation of biomass and toxin production in WLE CyanoHABs, we conducted in situ nutrient addition and 40% dilution microcosm bioassays in June and August 2019. During the June Sandusky Bay bloom, biomass production as well as hepatotoxic microcystin and neurotoxic anatoxin production were N and P co-limited with microcystin production becoming nutrient deplete under 40% dilution. During August, the Maumee Bay bloom produced microcystin under nutrient repletion with slight induced P limitation under 40% dilution, and the Sandusky Bay bloom produced anatoxin under N limitation in both dilution treatments. The results demonstrate the importance of nutrient limitation effects on microcystin and anatoxin production. To properly combat cyanotoxin and cyanobacterial biomass production in WLE, both N and P reduction efforts should be implemented in its watershed.

Elucidating controls on cyanobacteria bloom timing and intensity via Bayesian mechanistic modeling.

The adverse impacts of harmful algal blooms (HABs) are increasing worldwide. Lake Erie is a North American Great Lake highly affected by cultural eutrophication and summer cyanobacterial HABs. While phosphorus loading is a known driver of bloom size, more nuanced yet crucial questions remain. For example, it is unclear what mechanisms are primarily responsible for initiating cyanobacterial dominance and subsequent biomass accumulation. To address these questions, we develop a mechanistic model describing June-October dynamics of chlorophyll a, nitrogen, and phosphorus near the Maumee River outlet, where blooms typically initiate and are most severe. We calibrate the model to a new, geostatistically-derived dataset of daily water quality spanning 2008-2017. A Bayesian framework enables us to embed prior knowledge on system characteristics and test alternative model formulations. Overall, the best model formulation explains 42% of the variability in chlorophyll a and 83% of nitrogen, and better captures bloom timing than previous models. Our results, supported by cross validation, show that onset of the major midsummer bloom is associated with about a month of water temperatures above 20 °C (occurring 19 July to 6 August), consistent with when cyanobacteria dominance is usually reported. Decreased phytoplankton loss rate is the main factor enabling biomass accumulation, consistent with reduced zooplankton grazing on cyanobacteria. The model also shows that phosphorus limitation is most severe in August, and nitrogen limitation tends to occur in early autumn. Our results highlight the role of temperature in regulating bloom initiation and subsequent loss rates, and suggest that a 2 °C increase could lead to blooms that start about 10 days earlier and grow 23% more intense.

Isolation and Characterization of Rhizophydiales sp. (Chytridiomycota), Obligate Parasite of Planktothrix agardhii in a Laurentian Great Lakes Embayment.

Planktothrix agardhii dominates the cyanobacterial harmful algal bloom community in Sandusky Bay, Lake Erie (USA) from May through September. This filamentous cyanobacterium is host to a known obligate parasite; the chytrid Rhizophydium sp. During the 2018 bloom season, by utilizing dilution and single filament isolation techniques, 7 chytrid and 12 P. agardhii strains were isolated from Sandusky Bay. These 7 chytrids and a selection of P. agardhii hosts were then characterized with respect to infection rates. Infections by the isolated chytrids were specific to Planktothrix planktonic species and were not found on other filamentous cyanobacterial taxa present in the bay (Aphanizomenon sp. and Cuspidothrix sp.). Even among the potential P. agardhii host strains, individual chytrid isolates had different degrees of infectivity and showed preference for different host isolates, suggesting possible ecological partitioning even within the same sample population. Examining mechanisms of chytrid pathogenesis, the zoospores displayed a swarming pattern to attack and fracture the host filament and create new infection sites at the trichome termini. Infections by these parasitic chytrids also led to a release of intracellular microcystin toxins from the hosts. Additionally, infections were dependent on media type, highlighting the importance of media choice on experimental outcomes. Media in which chytrid swarming was observed closely matched the ionic strength of the natural environment. Understanding pathogenesis by fungal parasites will assist future efforts aimed at determining environmental factors favoring loss mechanisms for Planktothrix agardhii-dominated blooms.IMPORTANCE Whereas many studies have focused on the factors contributing to the establishment and persistence of cyanobacterial harmful algal blooms (cHABs), few studies have examined bloom pathogenesis. Chytrid fungi infect cyanobacteria and stimulate food web interactions through manipulation of previously hard to digest filaments and the release of nutrients to support heterotrophic microbes. Specifically, chytrids infective on filamentous Planktothrix agardhii exhibit a species-specific infection that fragments trichomes into shorter units that can be consumed more easily by grazers. Chytrid zoospores also serve as a high-quality food source for the lower food web. Understanding host-pathogen relationships and mechanisms of pathogenesis on cyanobacteria will be necessary to effectively model the ecology of cHABs.

Nutrient Capture from Aqueous Waste and Photocontrolled Fertilizer Delivery to Tomato Plants Using Fe(III)-Polysaccharide Hydrogels.

Inexpensive and sustainable methods are needed to reclaim nutrients from agricultural waste solutions for use as a fertilizer while decreasing nutrient runoff. Fe(III)-polysaccharide hydrogels are able to flocculate solids and absorb nutrients in liquid animal waste from Confined Animal Feeding Operations (CAFOs). Fe(III)-alginate beads absorbed 0.05 mg g-1 NH4 + and NO3 - from 100 ppm solutions at pH = 7, with > 80% phosphate uptake and ∼30% uptake of ammonium and nitrate. Ammonium uptake from a raw manure solution (1420 ppm NH4 +) showed a significant 0.7 mg g-1 uptake. Tomato plant trials carried out with Fe(III)-alginate hydrogel beads in greenhouse conditions showed controlled nutrient delivery for the plants compared to fertilizer solution with the same nutrient content. Plants showed an uptake of Fe from the gel beads, and Fe(III)-alginate hydrogel beads promoted root growth of the plants. The plants treated with nutrient-loaded Fe(III)-alginate hydrogels yielded comparable tomato harvest to plants treated with the conventional fertilizer solution.

Dissolved Microcystin Release Coincident with Lysis of a Bloom Dominated by Microcystis spp. in Western Lake Erie Attributed to a Novel Cyanophage.

Western Lake Erie (Laurentian Great Lakes) is prone to annual cyanobacterial harmful algal blooms (cHABs) dominated by Microcystis spp. that often yield microcystin toxin concentrations exceeding the federal EPA recreational contact advisory of 8 µg liter-1 In August 2014, microcystin levels were detected in finished drinking water above the World Health Organization 1.0 µg liter-1 threshold for consumption, leading to a 2-day disruption in the supply of drinking water for >400,000 residents of Toledo, Ohio (USA). Subsequent metatranscriptomic analysis of the 2014 bloom event provided evidence that release of toxin into the water supply was likely caused by cyanophage lysis that transformed a portion of the intracellular microcystin pool into the dissolved fraction, rendering it more difficult to eliminate during treatment. In August 2019, a similar increase in dissolved microcystins at the Toledo water intake was coincident with a viral lytic event caused by a phage consortium different in composition from what was detected following the 2014 Toledo water crisis. The most abundant viral sequence in metagenomic data sets was a scaffold from a putative member of the Siphoviridae, distinct from the Ma-LMM01-like Myoviridae that are typically documented to occur in western Lake Erie. This study provides further evidence that viral activity in western Lake Erie plays a significant role in transformation of microcystins from the particulate to the dissolved fraction and therefore requires monitoring efforts from local water treatment plants. Additionally, identification of multiple lytic cyanophages will enable the development of a quantitative PCR toolbox to assess viral activity during cHABs.IMPORTANCE Viral attack on cHABs may contribute to changes in community composition during blooms, as well as bloom decline, yet loss of bloom biomass does not eliminate the threat of cHAB toxicity. Rather, it may increase risks to the public by delivering a pool of dissolved toxin directly into water treatment utilities when the dominating Microcystis spp. are capable of producing microcystins. Detecting, characterizing, and quantifying the major cyanophages involved in lytic events will assist water treatment plant operators in making rapid decisions regarding the pool of microcystins entering the plant and the corresponding best practices to neutralize the toxin.

Toxic benthic freshwater cyanobacterial proliferations: Challenges and solutions for enhancing knowledge and improving monitoring and mitigation.

1. This review summarises knowledge on the ecology, toxin production, and impacts of toxic freshwater benthic cyanobacterial proliferations. It documents monitoring, management, and sampling strategies, and explores mitigation options. 2. Toxic proliferations of freshwater benthic cyanobacteria (taxa that grow attached to substrates) occur in streams, rivers, lakes, and thermal and meltwater ponds, and have been reported in 19 countries. Anatoxin- and microcystin-containing mats are most commonly reported (eight and 10 countries, respectively). 3. Studies exploring factors that promote toxic benthic cyanobacterial proliferations are limited to a few species and habitats. There is a hierarchy of importance in environmental and biological factors that regulate proliferations with variables such as flow (rivers), fine sediment deposition, nutrients, associated microbes, and grazing identified as key drivers. Regulating factors differ among colonisation, expansion, and dispersal phases. 4. New -omics-based approaches are providing novel insights into the physiological attributes of benthic cyanobacteria and the role of associated microorganisms in facilitating their proliferation. 5. Proliferations are commonly comprised of both toxic and non-toxic strains, and the relative proportion of these is the key factor contributing to the overall toxin content of each mat. 6. While these events are becoming more commonly reported globally, we currently lack standardised approaches to detect, monitor, and manage this emerging health issue. To solve these critical gaps, global collaborations are needed to facilitate the rapid transfer of knowledge and promote the development of standardised techniques that can be applied to diverse habitats and species, and ultimately lead to improved management.

Physical drivers facilitating a toxigenic cyanobacterial bloom in a major Great Lakes tributary.

The Maumee River is the primary source for nutrients fueling seasonal Microcystis-dominated blooms in western Lake Erie's open waters though such blooms in the river are infrequent. The river also serves as source water for multiple public water systems and a large food services facility in northwest Ohio, USA. On 20 September 2017, an unprecedented bloom was reported in the Maumee River estuary within the Toledo metropolitan area, which triggered a recreational water advisory. Here we (1) explore physical drivers likely contributing to the bloom's occurrence, and (2) describe the toxin concentration and bacterioplankton taxonomic composition. A historical analysis using ten-years of seasonal river discharge, water level, and local wind data identified two instances when high-retention conditions occurred over ≥10 days in the Maumee River estuary: in 2016 and during the 2017 bloom. Observation by remote sensing imagery supported the advection of cyanobacterial cells into the estuary from the lake during 2017 and the lack of an estuary bloom in 2016 due to a weak cyanobacterial bloom in the lake. A rapid-response survey during the 2017 bloom determined levels of the cyanotoxins, specifically microcystins, in excess of recreational contact limits at sites within the lower 20 km of the river while amplicon sequencing found these sites were dominated by Microcystis. These results highlight the need to broaden our understanding of physical drivers of cyanobacterial blooms within the interface between riverine and lacustrine systems, particularly as such blooms are expected to become more prominent in response to a changing climate.

Recent Patterns in Lake Erie Phosphorus and Chlorophyll a Concentrations in Response to Changing Loads.

Despite the initial success of extensive efforts to reduce phosphorus (P) loading to Lake Erie as a part of the Great Lakes Water Quality Agreement, Lake Erie appears to be undergoing a re-eutrophication and it is plagued by harmful algal blooms. To offer insights into potential lake responses under differing Maumee River loads and reveal recent changes with time, we explored patterns in phosphorus and chlorophyll a data from 2008 to 2018 collected in western Lake Erie near the mouth of the Maumee River. We found high, but relatively stable Maumee River and lake concentrations of total P (TP) and soluble reactive P (SRP) with no discernable annual or seasonal patterns. Maumee spring TP load was not strongly related to lake TP, and lake SRP concentrations were positively but weakly related to SRP loads. Lake TP was a strong predictor of chlorophyll a, but the relationship was weaker at sites closer to the Maumee. These results highlight spatial differences both in P concentration and the relationship between TP and chlorophyll a, and these indicate that spring phosphorus loads are a weak algal biomass predictor in the portion of the western basin of Lake Erie represented by these sampling stations.

Multiple cyanotoxin congeners produced by sub-dominant cyanobacterial taxa in riverine cyanobacterial and algal mats.

Benthic cyanobacterial proliferations in rivers are have been reported with increasing frequency worldwide. In the Eel and Russian rivers of California, more than a dozen dog deaths have been attributed to cyanotoxin toxicosis since 2000. Periphyton proliferations in these rivers comprise multiple cyanobacterial taxa capable of cyanotoxin production, hence there is uncertainty regarding which taxa are producing toxins. In this study, periphyton samples dominated by the cyanobacterial genera Anabaena spp. and Microcoleus spp. and the green alga Cladophora glomerata were collected from four sites in the Eel River catchment and one site in the Russian River. Samples were analysed for potential cyanotoxin producers using polymerase chain reaction (PCR) in concert with Sanger sequencing. Cyanotoxin concentrations were measured using liquid chromatography tandem-mass spectrometry, and anatoxin quota (the amount of cyanobacterial anatoxins per toxigenic cell) determined using droplet digital PCR. Sequencing indicated Microcoleus sp. and Nodularia sp. were the putative producers of cyanobacterial anatoxins and nodularins, respectively, regardless of the dominant taxa in the mat. Anatoxin concentrations in the mat samples varied from 0.1 to 18.6 µg g-1 and were significantly different among sites (p < 0.01, Wilcoxon test); however, anatoxin quotas were less variable (< 5-fold). Dihydroanatoxin-a was generally the most abundant variant in samples comprising 38% to 71% of the total anatoxins measured. Mats dominated by the green alga C. glomerata contained both anatoxins and nodularin-R at concentrations similar to those of cyanobacteria-dominated mats. This highlights that even when cyanobacteria are not the dominant taxa in periphyton, these mats may still pose a serious health risk and indicates that more widespread monitoring of all mats in a river are necessary.

A Multiplex Analysis of Potentially Toxic Cyanobacteria in Lake Winnipeg during the 2013 Bloom Season.

Lake Winnipeg (Manitoba, Canada), the world's 12th largest lake by area, is host to yearly cyanobacterial harmful algal blooms (cHABs) dominated by Aphanizomenon and Dolichospermum. cHABs in Lake Winnipeg are primarily a result of eutrophication but may be exacerbated by the recent introduction of dreissenid mussels. Through multiple methods to monitor the potential for toxin production in Lake Winnipeg in conjunction with environmental measures, this study defined the baseline composition of a Lake Winnipeg cHAB to measure potential changes because of dreissenid colonization. Surface water samples were collected in 2013 from 23 sites during summer and from 18 sites in fall. Genetic data and mass spectrometry cyanotoxin profiles identified microcystins (MC) as the most abundant cyanotoxin across all stations, with MC concentrations highest in the north basin. In the fall, mcyA genes were sequenced to determine which species had the potential to produce MCs, and 12 of the 18 sites were a mix of both Planktothrix and Microcystis. Current blooms in Lake Winnipeg produce low levels of MCs, but the capacity to produce cyanotoxins is widespread across both basins. If dreissenid mussels continue to colonize Lake Winnipeg, a shift in physicochemical properties of the lake because of faster water column clearance rates may yield more toxic blooms potentially dominated by microcystin producers.

Metatranscriptomic Analyses of Diel Metabolic Functions During a Microcystis Bloom in Western Lake Erie (United States).

This study examined diel shifts in metabolic functions of Microcystis spp. during a 48-h Lagrangian survey of a toxin-producing cyanobacterial bloom in western Lake Erie in the aftermath of the 2014 Toledo Water Crisis. Transcripts mapped to the genomes of recently sequenced lower Great Lakes Microcystis isolates showed distinct patterns of gene expression between samples collected across day (10:00 h, 16:00 h) and night (22:00 h, 04:00 h). Daytime transcripts were enriched in functions related to Photosystem II (e.g., psbA), nitrogen and phosphate acquisition, cell division (ftsHZ), heat shock response (dnaK, groEL), and uptake of inorganic carbon (rbc, bicA). Genes transcribed during nighttime included those involved in phycobilisome protein synthesis and Photosystem I core subunits. Hierarchical clustering and principal component analysis (PCA) showed a tightly clustered group of nighttime expressed genes, whereas daytime transcripts were separated from each other over the 48-h duration. Lack of uniform clustering within the daytime transcripts suggested that the partitioning of gene expression in Microcystis is dependent on both circadian regulation and physicochemical changes within the environment.

Science meets policy: A framework for determining impairment designation criteria for large waterbodies affected by cyanobacterial harmful algal blooms.

Toxic cyanobacterial harmful algal blooms (cyanoHABs) are one of the most significant threats to the security of Earth's surface freshwaters. In the United States, the Federal Water Pollution Control Act of 1972 (i.e., the Clean Water Act) requires that states report any waterbody that fails to meet applicable water quality standards. The problem is that for fresh waters impacted by cyanoHABs, no scientifically-based framework exists for making this designation. This study describes the development of a data-based framework using the Ohio waters of western Lake Erie as an exemplar for large lakes impacted by cyanoHABs. To address this designation for Ohio's open waters, the Ohio Environmental Protection Agency (EPA) assembled a group of academic, state and federal scientists to develop a framework that would determine the criteria for Ohio EPA to consider in deciding on a recreation use impairment designation due to cyanoHAB presence. Typically, the metrics are derived from on-lake monitoring programs, but for large, dynamic lakes such as Lake Erie, using criteria based on discrete samples is problematic. However, significant advances in remote sensing allows for the estimation of cyanoHAB biomass of an entire lake. Through multiple years of validation, we developed a framework to determine lake-specific criteria for designating a waterbody as impaired by cyanoHABs on an annual basis. While the criteria reported in this manuscript are specific to Ohio's open waters, the framework used to determine them can be applied to any large lake where long-term monitoring data and satellite imagery are available.

Reduced forms of nitrogen are a driver of non-nitrogen-fixing harmful cyanobacterial blooms and toxicity in Lake Erie.

Western Lake Erie (WLE) experiences anthropogenic eutrophication and annual, toxic cyanobacterial blooms of non-nitrogen (N) fixing Microcystis. Numerous studies have shown that bloom biomass is correlated with an increased proportion of soluble reactive phosphorus loading from the Maumee River. Long term monitoring shows that the proportion of the annual Maumee River N load of non-nitrate N, or total Kjeldahl nitrogen (TKN), has also increased significantly (Spearman's ρ = 0.68, p = 0.001) over the last few decades and is also significantly correlated to cyanobacterial bloom biomass (Spearman's ρ = 0.64, p = 0.003). The ratio of chemically reduced N to oxidized N (TKN:NO3) concentrations was also compared to extracted chlorophyll and phycocyanin concentrations from all weekly sampling stations within WLE from 2009 to 2015. Both chlorophyll (Spearman's ρ = 0.657, p < 0.0001) and phycocyanin (Spearman's ρ = 0.714, p < 0.0001) were significantly correlated with TKN:NO3. This correlation between the increasing fraction of chemically reduced N from the Maumee River and increasing bloom biomass demonstrates the urgent need to control N loading, in addition to current P load reductions, to WLE and similar systems impacted by non-N-fixing, toxin-producing cyanobacteria.

Establishing spatial and temporal patterns in Microcystis sediment seed stock viability and their relationship to subsequent bloom development in Western Lake Erie.

This study assessed the distribution, abundance, and viability of pre- and post-overwintering Microcystis sediment seed stocks in Western Lake Erie and how these variables are potentially related to past and subsequent bloom formation. We conducted a two-year spatiotemporal survey of vegetative seed stocks in Western Lake Erie, the region where annual algal blooms generally develop. Sediment was collected from 16 sites covering an area of 375 km2 and water column depths ranging from 3-9 meters. Sample collection occurred in November 2014, April 2015, November 2015, and April 2016. The abundance of total and potentially-toxic Microcystis cell equivalents were determined using quantitative polymerase chain reaction. A series of laboratory experiments using lake sediment were conducted to assess the viability of Microcystis vegetative seed stocks. Across all sampling periods, the abundance of total Microcystis in the sediment ranged from 6.6 x 10(4) to 1.7 x 10(9) cell equivalents g-1, and potentially-toxic Microcystis ranged from 1.4 x 10(3) to 4.7 x 10(6) cell equivalents g-1. The percent potentially-toxic Microcystis in the sediment ranged from <1% to 68% across all samples. Total Microcystis abundance diminished significantly over winter with densities in spring nearly 10 times less than the previous fall. However, despite cell loss from fall to spring, lab experiments demonstrated that remaining non-toxic and potentially-toxic cells were viable after the overwintering period. Further, lab grow-out experiments indicate that potentially-toxic strains recruited at a slightly higher rate than non-toxic strains, and may in part, contribute to the pattern of higher relative toxicity during early stages of the blooms. The abundance and distribution of overwintering cells did not correlate strongly to areas in the lake where subsequent summer blooms were most persistent. However, numerical analysis suggests that recruitment of benthic overwintering populations could help explain a portion of the initial rapid increase in bloom biomass and the spatial extent of this bloom initiation, particularly when recruitment is paired with subsequent growth in appropriate water column conditions.