Rapidly developed, optimized, and applied wastewater surveillance system for real-time monitoring of low-incidence, high-impact MPOX outbreak.

Recent MPOX viral resurgences have mobilized public health agencies around the world. Recognizing the significant risk of MPOX outbreaks, large-scale human testing, and immunization campaigns have been initiated by local, national, and global public health authorities. Recently, traditional clinical surveillance campaigns for MPOX have been complemented with wastewater surveillance (WWS), building on the effectiveness of existing wastewater programs that were built to monitor SARS-CoV-2 and recently expanded to include influenza and respiratory syncytial virus surveillance in wastewaters. In the present study, we demonstrate and further support the finding that MPOX viral fragments agglomerate in the wastewater solids fraction. Furthermore, this study demonstrates that the current, most commonly used MPOX assays are equally effective at detecting low titers of MPOX viral signal in wastewaters. Finally, MPOX WWS is shown to be more effective at passively tracking outbreaks and/or resurgences of the disease than clinical testing alone in smaller communities with low human clinical case counts of MPOX.

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.

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.

Environmental drivers of phytoplankton community dynamics in an agriculturally-influenced tributary in the lower Great Lakes.

Phytoplankton community composition in tributaries differs from that in their receiving waters, due to light limitation from suspended particles and other factors such as nutrient availability and temperature. This study was designed to manipulate light levels in early, mid, and late summer to determine the combined effects of light attenuation and naturally varying nutrient availability on phytoplankton community composition in an agriculturally-influenced tributary of the lower Great Lakes. In all trials, in situ microcosm experiments show that phytoplankton abundance increased under three light attenuation treatments (60 %, 75 %, and 85 % attenuation) relative to time-zero, but higher light attenuation reduced total phytoplankton abundance relative to controls. Highest phytoplankton diversity in terms of richness and evenness occurred in September (late summer), and across all three trials was lowest under the highest light attenuation treatments (85 %). Phytoplankton community composition followed a normal seasonal shift from diatoms dominating in June (early summer), followed by cyanobacteria dominating in mid to late summer. In general, lower light levels (especially 85 % attenuation) corresponded with an increased dominance of cyanobacteria. These findings support the hypothesis that phytoplankton abundance and diversity vary with light and nutrient availability and that light attenuation promotes the shift from buoyant cyanobacteria to other taxa more tolerant of low light levels.

Metatranscriptomic analysis reveals dissimilarity in viral community activity between an ice-free and ice-covered winter in Lake Erie.

Winter is a relatively under-studied season in freshwater ecology. The paucity of wintertime surveys has led to a lack of knowledge regarding microbial community activity during the winter in Lake Erie, a North American Great Lake. Viruses shape microbial communities and regulate biogeochemical cycles by acting as top-down controls, yet very few efforts have been made to examine active virus populations during the winter in Lake Erie. Furthermore, climate change-driven declines in seasonal ice cover have been shown to influence microbial community structure, but no studies have compared viral community activity between different ice cover conditions. We surveyed surface water metatranscriptomes for viral hallmark genes as a proxy for active virus populations and compared activity metrics between ice-covered and ice-free conditions from two sampled winters. Transcriptionally active viral communities were detected in both winters, spanning diverse phylogenetic clades of putative bacteriophage (Caudoviricetes), giant viruses (Nucleocytoviricota, or NCLDV), and RNA viruses (Orthornavirae). However, viral community activity metrics revealed pronounced differences between the ice-covered and ice-free winters. Viral community composition was distinct between winters and viral hallmark gene richness was reduced in the ice-covered relative to the ice-free conditions. In addition, the observed differences in viral communities correlated with microbial community activity metrics. Overall, these findings contribute to our understanding of the viral populations that are active during the winter in Lake Erie and suggest that viral community activity may be associated with ice cover extent.IMPORTANCEAs seasonal ice cover is projected to become increasingly rare on large temperate lakes, there is a need to understand how microbial communities might respond to changing ice conditions. Although it is widely recognized that viruses impact microbial community structure and function, there is little known regarding wintertime viral activity or the relationship between viral activity and ice cover extent. Our metatranscriptomic analyses indicated that viruses were transcriptionally active in the winter surface waters of Lake Erie. These findings also expanded the known diversity of viral lineages in the Great Lakes. Notably, viral community activity metrics were significantly different between the two sampled winters. The pronounced differences we observed in active viral communities between the ice-covered and ice-free samples merit further research regarding how viral communities will function in future, potentially ice-free, freshwater systems.

Lake Erie ice is a repository of organisms.

Organism abundance and diversity were assessed in Lake Erie ice samples using sequences derived from a combined metagenomic and metatranscriptomic analysis. The 68,417 unique sequences were from Bacteria (77.5%), Eukarya (22.3%), and Archaea (0.2%) and indicated diverse species of organisms from 32 bacterial, 8 eukaryotic, and 2 archaeal taxonomic groups.

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.

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.

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.

Evaluating and optimizing Acid-pH and Direct Lysis RNA extraction for SARS-CoV-2 RNA detection in whole saliva.

COVID-19 has been a global public health and economic challenge. Screening for the SARS-CoV-2 virus has been a key part of disease mitigation while the world continues to move forward, and lessons learned will benefit disease detection beyond COVID-19. Saliva specimen collection offers a less invasive, time- and cost-effective alternative to standard nasopharyngeal swabs. We optimized two different methods of saliva sample processing for RT-qPCR testing. Two methods were optimized to provide two cost-efficient ways to do testing for a minimum of four samples by pooling in a 2.0 mL tube and decrease the need for more highly trained personnel. Acid-pH-based RNA extraction method can be done without the need for expensive kits. Direct Lysis is a quick one-step reaction that can be applied quickly. Our optimized Acid-pH and Direct Lysis protocols are reliable and reproducible, detecting the beta-2 microglobulin (B2M) mRNA in saliva as an internal control from 97 to 96.7% of samples, respectively. The cycle threshold (Ct) values for B2M were significantly higher in the Direct Lysis protocol than in the Acid-pH protocol. The limit of detection for N1 gene was higher in Direct Lysis at ≤ 5 copies/µL than Acid-pH. Saliva samples collected over the course of several days from two COVID-positive individuals demonstrated Ct values for N1 that were consistently higher from Direct Lysis compared to Acid-pH. Collectively, this work supports that each of these techniques can be used to screen for SARS-CoV-2 in saliva for a cost-effective screening platform.

SARS-CoV-2 viral titer measurements in Ontario, Canada wastewaters throughout the COVID-19 pandemic.

During the COVID-19 pandemic, the Province of Ontario, Canada, launched a wastewater surveillance program to monitor SARS-CoV-2, inspired by the early work and successful forecasts of COVID-19 waves in the city of Ottawa, Ontario. This manuscript presents a dataset from January 1, 2021, to March 31, 2023, with RT-qPCR results for SARS-CoV-2 genes and PMMoV from 107 sites across all 34 public health units in Ontario, covering 72% of the province's and 26.2% of Canada's population. Sampling occurred 2-7 times weekly, including geographical coordinates, serviced populations, physico-chemical water characteristics, and flowrates. In doing so, this manuscript ensures data availability and metadata preservation to support future research and epidemic preparedness through detailed analyses and modeling. The dataset has been crucial for public health in tracking disease locally, especially with the rise of the Omicron variant and the decline in clinical testing, highlighting wastewater-based surveillance's role in estimating disease incidence in Ontario.

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.

A combined microscopy and single-cell sequencing approach reveals the ecology, morphology, and phylogeny of uncultured lineages of zoosporic fungi.

Environmental DNA analyses of fungal communities typically reveal a much larger diversity than can be ascribed to known species. Much of this hidden diversity lies within undescribed fungal lineages, especially the early diverging fungi (EDF). Although these EDF often represent new lineages even at the phylum level, they have never been cultured, making their morphology and ecology uncertain. One of the methods to characterize these uncultured fungi is a single-cell DNA sequencing approach. In this study, we established a large data set of single-cell sequences of EDF by manually isolating and photographing parasitic fungi on various hosts such as algae, protists, and micro-invertebrates, combined with subsequent long-read sequencing of the ribosomal DNA locus (rDNA). We successfully obtained rDNA sequences of 127 parasitic fungal cells, which clustered into 71 phylogenetic lineages belonging to seven phylum-level clades of EDF: Blastocladiomycota, Chytridiomycota, Aphelidiomycota, Rozellomycota, and three unknown phylum-level clades. Most of our single cells yielded novel sequences distinguished from both described taxa and existing metabarcoding data, indicating an expansive and hidden diversity of parasitic taxa of EDF. We also revealed an unexpected diversity of endobiotic Olpidium-like chytrids and hyper-parasitic lineages. Overall, by combining photographs of parasitic fungi with phylogenetic analyses, we were able to better understand the ecological function and morphology of many of the branches on the fungal tree of life known only from DNA sequences. IMPORTANCE Much of the diversity of microbes from natural habitats, such as soil and freshwater, comprise species and lineages that have never been isolated into pure culture. In part, this stems from a bias of culturing in favor of saprotrophic microbes over the myriad symbiotic ones that include parasitic and mutualistic relationships with other taxa. In the present study, we aimed to shed light on the ecological function and morphology of the many undescribed lineages of aquatic fungi by individually isolating and sequencing molecular barcodes from 127 cells of host-associated fungi using single-cell sequencing. By adding these sequences and their photographs into the fungal tree, we were able to understand the morphology of reproductive and vegetative structures of these novel fungi and to provide a hypothesized ecological function for them. These individual host-fungal cells revealed themselves to be complex environments despite their small size; numerous samples were hyper-parasitized with other zoosporic fungal lineages such as Rozellomycota.

Actionable wastewater surveillance: application to a university residence hall during the transition between Delta and Omicron resurgences of COVID-19.

Wastewater surveillance has gained traction during the COVID-19 pandemic as an effective and non-biased means to track community infection. While most surveillance relies on samples collected at municipal wastewater treatment plants, surveillance is more actionable when samples are collected "upstream" where mitigation of transmission is tractable. This report describes the results of wastewater surveillance for SARS-CoV-2 at residence halls on a university campus aimed at preventing outbreak escalation by mitigating community spread. Another goal was to estimate fecal shedding rates of SARS-CoV-2 in a non-clinical setting. Passive sampling devices were deployed in sewer laterals originating from residence halls at a frequency of twice weekly during fall 2021 as the Delta variant of concern continued to circulate across North America. A positive detection as part of routine sampling in late November 2021 triggered daily monitoring and further isolated the signal to a single wing of one residence hall. Detection of SARS-CoV-2 within the wastewater over a period of 3 consecutive days led to a coordinated rapid antigen testing campaign targeting the residence hall occupants and the identification and isolation of infected individuals. With knowledge of the number of individuals testing positive for COVID-19, fecal shedding rates were estimated to range from 3.70 log10 gc ‧ g feces-1 to 5.94 log10 gc ‧ g feces-1. These results reinforce the efficacy of wastewater surveillance as an early indicator of infection in congregate living settings. Detections can trigger public health measures ranging from enhanced communications to targeted coordinated testing and quarantine.

Microcystin aids in cold temperature acclimation: Differences between a toxic Microcystis wildtype and non-toxic mutant.

For Microcystis aeruginosa PCC 7806, temperature decreases from 26 °C to 19 °C double the microcystin quota per cell during growth in continuous culture. Here we tested whether this increase in microcystin provided M. aeruginosa PCC 7806 with a fitness advantage during colder-temperature growth by comparing cell concentration, cellular physiology, reactive oxygen species damage, and the transcriptomics-inferred metabolism to a non-toxigenic mutant strain M. aeruginosa PCC 7806 ΔmcyB. Photo-physiological data combined with transcriptomic data revealed metabolic changes in the mutant strain during growth at 19 °C, which included increased electron sinks and non-photochemical quenching. Increased gene expression was observed for a glutathione-dependent peroxiredoxin during cold treatment, suggesting compensatory mechanisms to defend against reactive oxygen species are employed in the absence of microcystin in the mutant. Our observations highlight the potential selective advantages of a longer-term defensive strategy in management of oxidative stress (i.e., making microcystin) vs the shorter-term proactive strategy of producing cellular components to actively dissipate or degrade oxidative stress agents.

Cool temperature acclimation in toxigenic Microcystis aeruginosa PCC 7806 and its non-toxigenic mutant.

For Microcystis aeruginosa PCC 7806, temperature decreases from 26° C to 19° C double the microcystin quota per cell during growth in continuous culture. Here we tested whether this increase in microcystin provided M. aeruginosa PCC 7806 with a fitness advantage during colder-temperature growth by comparing cell concentration, cellular physiology, and the transcriptomics-inferred metabolism to a non-toxigenic mutant strain M. aeruginosa PCC 7806 ΔmcyB. Photo-physiological data combined with transcriptomic data revealed metabolic changes in the mutant strain during growth at 19° C, which included increased electron sinks and non-photochemical quenching. Increased gene expression was observed for a glutathione-dependent peroxiredoxin during cold treatment, suggesting compensatory mechanisms to defend against reactive oxygen species are employed in the absence of microcystin in the mutant. Our observations highlight the potential selective advantages of a longer-term defensive strategy in management of oxidative stress (i.e., making microcystin) vs the shorter-term proactive strategy of producing cellular components to actively dissipate or degrade oxidative stress agents.

Genomic comparison of Planktothrix agardhii isolates from a Lake Erie embayment.

Planktothrix agardhii is a filamentous cyanobacterial species that dominates harmful algal blooms in Sandusky Bay, Lake Erie and other freshwater basins across the world. P. agardhii isolates were obtained from early (June) blooms via single filament isolation; eight have been characterized from 2016, and 12 additional isolates have been characterized from 2018 for a total of 20 new cultures. These novel isolates were processed for genomic sequencing, where reads were used to generate scaffolds and contigs which were annotated with DIAMOND BLAST hit, Pfam, and GO. Analyses include whole genome alignment to generate phylogenetic trees and comparison of genetic rearrangements between isolates. Nitrogen acquisition and metabolism was compared across isolates. Secondary metabolite production was genetically explored including microcystins, two types of aeruginosin clusters, anabaenopeptins, cyanopeptolins, microviridins, and prenylagaramides. Two common and 4 unique CRISPR-cas islands were analyzed for similar sequences across all isolates and against the known Planktothrix-specific cyanophage, PaV-LD. Overall, the uniqueness of each genome from Planktothrix blooms sampled from the same site and at similar times belies the unexplored diversity of this genus.