An assessment of a biosensor system for the quantification of microcystins in freshwater cyanobacterial blooms.

Microcystin-producing cyanobacterial blooms are a global issue threatening drinking water supplies and recreation on lakes and beaches. Direct measurement of microcystins is the only way to ensure waters have concentrations below guideline concentrations; however, analyzing water for microcystins takes several hours to days to obtain data. We tested LightDeck Diagnostics' bead beater cell lysis and two versions of the quantification system designed to give microcystin concentrations within 20 min and compared it to the standard freeze-thaw cycle lysis method and ELISA quantification. The bead beater lyser was only 30 % effective at extracting microcystins compared to freeze-thaw. When considering freeze-thaw samples analyzed in 2021, there was good agreement between ELISA and LightDeck version 2 (n = 152; R2 = 0.868), but the LightDeck slightly underestimated microcystins (slope of 0.862). However, we found poor relationships between LightDeck version 2 and ELISA in 2022 (n = 49, slopes 0.60 to 1.6; R2 < 0.6) and LightDeck version 1 (slope = 1.77 but also a high number of less than quantifiable concentrations). After the quantification issues are resolved, combining the LightDeck system with an already-proven rapid lysis method (such as microwaving) will allow beach managers and water treatment operators to make quicker, well-informed decisions.

A tale of two blooms: do ecological paradigms for algal bloom success and succession require revisiting?

Lake Erie algal bloom discussions have historically focused on cyanobacteria, with foundational "blooms like it hot" and "high nutrient" paradigms considered as primary drivers behind cyanobacterial bloom success. Yet, recent surveys have rediscovered winter-spring diatom blooms, introducing another key player in the Lake Erie eutrophication and algal bloom story which has been historically overlooked. These blooms (summer vs. winter) have been treated as solitary events separated by spatial and temporal gradients. However, new evidence suggests they may not be so isolated, linked in a manner that manifests as an algal bloom cycle. Equally notable are the emerging reports of cyanobacterial blooms in cold and/or oligotrophic freshwaters, which have been interpreted by some as shifts in classical bloom paradigms. These emerging bloom reports have led many to ask "what is a bloom?". Furthermore, questioning classic paradigms has caused others to wonder if we are overlooking additional factors which constrain bloom success. In light of emerging data and ideas, we revisited foundational concepts within the context of Lake Erie algal blooms and derived five key take-aways: 1) Additional bloom-formers (diatoms) need to be included in Lake Erie algal discussions, 2) The term "bloom" must be reinforced with a clear definition and quantitative metrics for each event, 3) Algal blooms should not be studied solitarily, 4) Shifts in physiochemical conditions serve as an alternative interpretation to potential shifts in ecological paradigms, 5) Additional factors which constrain bloom success and succession (i.e., pH and light) require consideration.

Spatio-temporal Trends of Mercury and Stable Isotopes in Lower Food Web of Winam Gulf, Lake Victoria.

Components of the lower food web (mussels, Caridina and Omena) were collected from stations from Winam Gulf, Lake Victoria, Kenya in 2022 and 2023 to analyze for stable isotopes and total mercury (THg). Temporal comparisons were made with data generated for the same species in 1998. Values of δ15N in mussels and Caridina were similar (6.89‰ vs. 6.78 ± 0.13‰), while Omena occupied an elevated trophic position (9.97 ± 0.24‰) with minor shifts in δ15N over time. All species had elevated δ13C values in 2022-2023 versus 1998 supportive of enhanced eutrophication in the Gulf. THg concentrations exhibited modest spatial differences between sites (< 2.6 fold), but not between Caridina and Omena. Larger temporal differences were apparent relative to spatial patterns with THg concentrations decreasing in study species by 2.8 to 4.1-fold between years. An exposure assessment indicated that Omena, commonly found in local markets, can be consumed up to 0.74 kg/month without generating excess THg exposures.

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.

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.

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.

Terrestrial Origin for Abundant Riverine Nanoscale Ice-Nucleating Particles.

Ice-nucleating particles (INPs) associated with fresh waters are a neglected, but integral component of the water cycle. Abundant INPs were identified from surface waters of both the Maumee River and Lake Erie with ice nucleus spectra spanning a temperature range from -3 to -15 °C. The majority of river INPs were submicron in size and attributed to biogenic macromolecules, inferred from the denaturation of ice-nucleation activity by heat. In a watershed dominated by row-crop agriculture, higher concentrations of INPs were found in river samples compared to lake samples. Further, ice-nucleating temperatures differed between river and lake samples, which indicated different populations of INPs. Seasonal analysis of INPs that were active at warmer temperatures (≥-10 °C; INP-10) showed their concentration to correlate with river discharge, suggesting a watershed origin of these INPs. A terrestrial origin for INPs in the Maumee River was further supported by a correspondence between the ice-nucleation signatures of river INPs and INPs derived from the soil fungus Mortierella alpina. Aerosols derived from turbulence features in the river carry INP-10, although their potential influence on regional weather is unclear. INP-10 contained within aerosols generated from a weir spanning the river, ranged in concentration from 1 to 11 INP m-3, which represented a fold-change of 3.2 over average INP-10 concentrations sampled from aerosols at control locations.

Ecophysiological Examination of the Lake Erie Microcystis Bloom in 2014: Linkages between Biology and the Water Supply Shutdown of Toledo, OH.

Annual cyanobacterial blooms dominated by Microcystis have occurred in western Lake Erie (U.S./Canada) during summer months since 1995. The production of toxins by bloom-forming cyanobacteria can lead to drinking water crises, such as the one experienced by the city of Toledo in August of 2014, when the city was rendered without drinking water for >2 days. It is important to understand the conditions and environmental cues that were driving this specific bloom to provide a scientific framework for management of future bloom events. To this end, samples were collected and metatranscriptomes generated coincident with the collection of environmental metrics for eight sites located in the western basin of Lake Erie, including a station proximal to the water intake for the city of Toledo. These data were used to generate a basin-wide ecophysiological fingerprint of Lake Erie Microcystis populations in August 2014 for comparison to previous bloom communities. Our observations and analyses indicate that, at the time of sample collection, Microcystis populations were under dual nitrogen (N) and phosphorus (P) stress, as genes involved in scavenging of these nutrients were being actively transcribed. Targeted analysis of urea transport and hydrolysis suggests a potentially important role for exogenous urea as a nitrogen source during the 2014 event. Finally, simulation data suggest a wind event caused microcystin-rich water from Maumee Bay to be transported east along the southern shoreline past the Toledo water intake. Coupled with a significant cyanophage infection, these results reveal that a combination of biological and environmental factors led to the disruption of the Toledo water supply. This scenario was not atypical of reoccurring Lake Erie blooms and thus may reoccur in the future.

Effects of increasing nitrogen and phosphorus concentrations on phytoplankton community growth and toxicity during Planktothrix blooms in Sandusky Bay, Lake Erie.

Sandusky Bay experiences annual toxic cyanobacterial blooms dominated by Planktothrix agardhii/suspensa. To further understand the environmental drivers of these events, we evaluated changes in the growth response and toxicity of the Planktothrix-dominated blooms to nutrient amendments with orthophosphate (PO4) and inorganic and organic forms of dissolved nitrogen (N; ammonium (NH4), nitrate (NO3) and urea) over the bloom season (June - October). We complemented these with a metagenomic analysis of the planktonic microbial community. Our results showed that bloom growth and microcystin (MC) concentrations responded more frequently to additions of dissolved N than PO4, and that the dual addition of NH4 + PO4 and Urea + PO4 yielded the highest MC concentrations in 54% of experiments. Metagenomic analysis confirmed that P. agardhii/suspensa was the primary MC producer. The phylogenetic distribution of nifH revealed that both heterocystous cyanobacteria and heterotrophic proteobacteria had the genetic potential for N2 fixation in Sandusky Bay. These results suggest that as best management practices are developed for P reductions in Sandusky Bay, managers must be aware of the negative implications of not managing N loading into this system as N may significantly impact cyanobacterial bloom size and toxicity.

Draft genome sequences of three Planktothrix rubescens strains cultivated from a eutrophic lake in Norwalk, Ohio (USA).

Draft genomes were generated for three filamentous toxin-producing cyanobacterial strains cultivated from aquatic sources in Ohio sequenced by NovaSeq S4. Here, we report the classification and genome statistics of Planktothrix rubescens PR221, PR222, and PR223.

Metagenome-assembled genome sequences of two cyanobacterial cultures from Homa Bay County, Kenya.

Metagenome-assembled genomes were generated for two xenic cyanobacterial strains collected from aquatic sources in Kenya and sequenced by NovaSeq S4. Here, we report the classification and genome statistics of Microcystis panniformis WG22 and Limnospira fusiformis LS22.

The recent disappearance of a persistent Planktothrix bloom: Characterization of a regime shift in the phytoplankton of Sandusky Bay (USA).

Sandusky Bay is the drowned mouth of the Sandusky River in the southwestern portion of Lake Erie. The bay is a popular recreation location and a regional source for drinking water. Like the western basin of Lake Erie, Sandusky Bay is known for being host to summer cyanobacterial harmful algal blooms (cHABs) year after year, fueled by runoff from the predominantly agricultural watershed and internal loading of legacy nutrients (primarily phosphorus). Since at least 2003, Sandusky Bay has harbored a microcystin-producing bloom of Planktothrix agardhii, a species of filamentous cyanobacteria that thrives in low light conditions. Long-term sampling (2003-2018) of Sandusky Bay revealed regular Planktothrix-dominated blooms during the summer months, but in recent years (2019-2022), 16S rRNA gene community profiling revealed that Planktothrix has largely disappeared. From 2017-2022, microcystin decreased well below the World Health Organization (WHO) guidelines. Spring TN:TP ratios increased in years following dam removal, yet there were no statistically significant shifts in other physicochemical variables, such as water temperature and water clarity. With the exception of the high bloom of Planktothrix in 2018, there was no statistical difference in chlorophyll during all other years. Concurrent with the disappearance of Planktothrix, Cyanobium spp. have become the dominant cyanobacterial group. The appearance of other potential toxigenic genera (i.e., Aphanizomenon, Dolichospermum, Cylindrospermopsis) may motivate monitoring of new toxins of concern in Sandusky Bay. Here, we document the regime shift in the cyanobacterial community and propose evidence supporting the hypothesis that the decline in the Planktothrix bloom was linked to the removal of an upstream dam on the Sandusky River.

Increasing mercury bioaccumulation and biomagnification rates of Nile perch (Lates niloticus L.) in Winam Gulf, Lake Victoria, Kenya.

The Nile perch (Lates niloticus L.) commercial fishery for Lake Victoria in East Africa is an important source of revenue and employment. We focused on shifts in food web structure and total mercury (THg) bioaccumulation and biomagnification in Nile perch, and lower food web items collected from Winam Gulf (Kenya) sampled 24 years apart (1998 and 2022). Stable isotope carbon (δ13C) values were higher in all species from 2022 compared to 1998. Stable nitrogen isotope (δ15N) values in baseline organisms were lower in 2022 compared to 1998. In Nile perch, δ15N values were correlated with total length, but the δ15N-length regressions were steeper in 1998 compared to 2022 except for one large (158 cm) Nile perch from 1998 with an uncharacteristically low δ15N value. Total Hg concentrations were lower in lower trophic species from 2022 compared to 1998. However, the THg bioaccumulation rate (as a function of fish length) in Nile perch was greater in 2022 compared to 1998 resulting in 24.2 % to 42.4 % higher wet weight dorsal THg concentrations in 2022 Nile perch for market slot size (50 to 85 cm) fish. The contrasting observations of increased THg bioaccumulation with size in 2022 against decreases in the rate of trophic increase with size and lower THg concentrations of lower food web items imply reduced fish growth and potential bioenergetic stressors on Winam Gulf Nile perch. All samples except 1 large Nile perch (139 cm total length collected in 2022) had THg concentrations below the European Union trade limit (500 ng/g wet weight). However, for more vulnerable individuals (women, children and frequent fish eaters), we recommend a decrease in maximum monthly meal consumption for 55-75 cm Nile perch from 16 meals per month calculated for 1998 to a limit of 8 meals per month calculated for 2022.

Bacterial community and cyanotoxin gene distribution of the Winam Gulf, Lake Victoria, Kenya.

The Winam Gulf (Kenya) is frequently impaired by cyanobacterial harmful algal blooms (cHABs) due to inadequate wastewater treatment and excess agricultural nutrient input. While phytoplankton in Lake Victoria have been characterized using morphological criteria, our aim is to identify potential toxin-producing cyanobacteria using molecular approaches. The Gulf was sampled over two successive summer seasons, and 16S and 18S ribosomal RNA gene sequencing was performed. Additionally, key genes involved in production of cyanotoxins were examined by quantitative PCR. Bacterial communities were spatially variable, forming distinct clusters in line with regions of the Gulf. Taxa associated with diazotrophy were dominant near Homa Bay. On the eastern side, samples exhibited elevated cyrA abundances, indicating genetic capability of cylindrospermopsin synthesis. Indeed, near the Nyando River mouth in 2022, cyrA exceeded 10 million copies L-1 where there were more than 6000 Cylindrospermopsis spp. cells mL-1. In contrast, the southwestern region had elevated mcyE gene (microcystin synthesis) detections near Homa Bay where Microcystis and Dolichospermum spp. were observed. These findings show that within a relatively small embayment, composition and toxin synthesis potential of cHABs can vary dramatically. This underscores the need for multifaceted management approaches and frequent cyanotoxin monitoring to reduce human health impacts.

Declines in ice cover are accompanied by light limitation responses and community change in freshwater diatoms.

The rediscovery of diatom blooms embedded within and beneath the Lake Erie ice cover (2007-2012) ignited interest in psychrophilic adaptations and winter limnology. Subsequent studies determined the vital role ice plays in winter diatom ecophysiology as diatoms partition to the underside of ice, thereby fixing their location within the photic zone. Yet, climate change has led to widespread ice decline across the Great Lakes, with Lake Erie presenting a nearly "ice-free" state in several recent winters. It has been hypothesized that the resultant turbid, isothermal water column induces light limitation amongst winter diatoms and thus serves as a competitive disadvantage. To investigate this hypothesis, we conducted a physiochemical and metatranscriptomic survey that spanned spatial, temporal, and climatic gradients of the winter Lake Erie water column (2019-2020). Our results suggest that ice-free conditions decreased planktonic diatom bloom magnitude and altered diatom community composition. Diatoms increased their expression of various photosynthetic genes and iron transporters, which suggests that the diatoms are attempting to increase their quantity of photosystems and light-harvesting components (a well-defined indicator of light limitation). We identified two gene families which serve to increase diatom fitness in the turbid ice-free water column: proton-pumping rhodopsins (a potential second means of light-driven energy acquisition) and fasciclins (a means to "raft" together to increase buoyancy and co-locate to the surface to optimize light acquisition). With large-scale climatic changes already underway, our observations provide insight into how diatoms respond to the dynamic ice conditions of today and shed light on how they will fare in a climatically altered tomorrow.

Effects of water movement and temperature on Rhizophydium infection of Planktothrix in a shallow hypereutrophic lake.

Grand Lake St. Marys (GLSM) is a popular recreational lake located in western Ohio, United States, generating nearly $150 million in annual revenue. However, recurring algal blooms dominated by Planktothrix agardhii, which can produce harmful microcystin toxins, have raised concerns about water safety and negatively impacted the local economy. Planktothrix agardhii is host to a number of parasites and pathogens, including an obligate fungal parasite in the Chytridiomycota (chytrids). In this study, we investigated the potential of these chytrid (Rhizophydium sp.) to infect P. agardhii blooms in the environment by modifying certain environmental conditions thought to limit infection prevalence in the wild. With a focus on temperature and water mixing, mesocosms were designed to either increase or decrease water flow compared to the control (water outside the mesocosm). In the control and water circulation mesocosms, infections were found infrequently and were found on less than 0.75% of the Planktothrix population. On the other hand, by decreasing the water flow to stagnation, chytrid infections were more frequent (found in nearly 3x as many samples) and more prevalent, reaching a maximum infection rate of 4.12%. In addition, qPCR coupled with 16S-18S sequencing was utilized to confirm the genetic presence of both host and parasite, as well as to better understand the effect of water circulation on the community composition. Statistical analysis of the data confirmed that chytrid infection was dependent on water temperature, with infections predominantly occurring between 19°C and 23°C. Additionally, water turbulence can significantly reduce the infectivity of chytrids, as infections were mostly found in stagnant mesocosms. Further, decreasing the water circulation promoted the growth of the cyanobacterial population, while increasing water agitation promoted the growth of green algae (Chlorophyta). This study starts to explore the environmental factors that affect chytrid pathogenesis which can provide valuable insights into controlling measures to reduce the prevalence of harmful algal blooms and improve water quality in GLSM and similarly affected waterbodies.

Interactions between chytrids cause variable infection strategies on harmful algal bloom forming species.

Cyanobacteria have a great diversity of natural enemies, such as herbivores and pathogens, including fungal pathogens within the Chytridiomycota (chytrids). While these pathogens have been previously described on a select number of cyanobacterial hosts and are suspected to play a significant ecological role, little is understood about species interactions and how competition between parasites can affect epidemic development and bloom formation. Here, three Planktothrix agardhii isolates from Sandusky Bay, Lake Erie (OH, USA) were challenged in monoculture and polyculture against infection by three isolates (C1, C2, C10) of their obligate chytrid fungal pathogen, Rhizophydiales sp. The chytrid isolates were inoculated as single isolates or a mixture of up to three different isolates. In monoculture, host isolates were characterized as highly susceptible (P. agardhii 1030), moderately susceptible (P. agardhii 1808) or mostly resistant (P. agardhii 1801). Co-infection of chytrid isolates on the highly susceptible host isolate had an additive effect on chytrid prevalence, leading to a culture crash where 2 or 3 chytrid isolates were present. Co-infection of chytrid isolates on the moderately susceptible and mostly resistant isolates had no effect on chytrid infection outcome or prevalence compared to infection with a single isolate. In polyculture, the effect on host growth was most significant in the single chytrid isolate treatment, which was attenuated with the addition of mixed chytrid treatments. Genetic analysis of the resulting population after the experimental period showed a tendency for the chytrid isolate C1 and P. agardhii 1801 to dominate in mixed population samples. Two different interspecific interactions seem to be in play; varied parasite infection strategies allow for the amplification of infection prevalence due to mixed chytrids in a susceptible monoculture, or competition allows for the dominance of a single chytrid isolate in monoculture and the reduction of infection prevalence in a host polyculture. This work thus highlights how interactions between chytrid infections can change the course of epidemic development and harmful algal bloom formation.

Climate change and the aquatic continuum: A cyanobacterial comeback story.

Billions of years ago, the Earth's waters were dominated by cyanobacteria. These microbes amassed to such formidable numbers, they ushered in a new era-starting with the Great Oxidation Event-fuelled by oxygenic photosynthesis. Throughout the following eon, cyanobacteria ceded portions of their global aerobic power to new photoautotrophs with the rise of eukaryotes (i.e. algae and higher plants), which co-existed with cyanobacteria in aquatic ecosystems. Yet while cyanobacteria's ecological success story is one of the most notorious within our planet's biogeochemical history, scientists to this day still seek to unlock the secrets of their triumph. Now, the Anthropocene has ushered in a new era fuelled by excessive nutrient inputs and greenhouse gas emissions, which are again reshaping the Earth's biomes. In response, we are experiencing an increase in global cyanobacterial bloom distribution, duration, and frequency, leading to unbalanced, and in many instances degraded, ecosystems. A critical component of the cyanobacterial resurgence is the freshwater-marine continuum: which serves to transport blooms, and the toxins they produce, on the premise that "water flows downhill". Here, we identify drivers contributing to the cyanobacterial comeback and discuss future implications in the context of environmental and human health along the aquatic continuum. This Minireview addresses the overlooked problem of the freshwater to marine continuum and the effects of nutrients and toxic cyanobacterial blooms moving along these waters. Marine and freshwater research have historically been conducted in isolation and independently of one another. Yet, this approach fails to account for the interchangeable transit of nutrients and biology through and between these freshwater and marine systems, a phenomenon that is becoming a major problem around the globe. This Minireview highlights what we know and the challenges that lie ahead.

Multi-year molecular quantification and 'omics analysis of Planktothrix-specific cyanophage sequences from Sandusky Bay, Lake Erie.

Introduction: Planktothrix agardhii is a microcystin-producing cyanobacterium found in Sandusky Bay, a shallow and turbid embayment of Lake Erie. Previous work in other systems has indicated that cyanophages are an important natural control factor of harmful algal blooms. Currently, there are few cyanophages that are known to infect P. agardhii, with the best-known being PaV-LD, a tail-less cyanophage isolated from Lake Donghu, China. Presented here is a molecular characterization of Planktothrix specific cyanophages in Sandusky Bay. Methods and Results: Putative Planktothrix-specific viral sequences from metagenomic data from the bay in 2013, 2018, and 2019 were identified by two approaches: homology to known phage PaV-LD, or through matching CRISPR spacer sequences with Planktothrix host genomes. Several contigs were identified as having viral signatures, either related to PaV-LD or potentially novel sequences. Transcriptomic data from 2015, 2018, and 2019 were also employed for the further identification of cyanophages, as well as gene expression of select viral sequences. Finally, viral quantification was tested using qPCR in 2015-2019 for PaV-LD like cyanophages to identify the relationship between presence and gene expression of these cyanophages. Notably, while PaV-LD like cyanophages were in high abundance over the course of multiple years (qPCR), transcriptomic analysis revealed only low levels of viral gene expression. Discussion: This work aims to provide a broader understanding of Planktothrix cyanophage diversity with the goals of teasing apart the role of cyanophages in the control and regulation of harmful algal blooms and designing monitoring methodology for potential toxin-releasing lysis events.