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1. Metagenomic and Metatranscriptomic Insights into Population Diversity of Microcystis Blooms: Spatial and Temporal Dynamics of mcy Genotypes, Including a Partial Operon That Can Be Abundant and Expressed

   https://doi.org/10.1128/aem.02464-21  

   Yancey, Colleen E. ; Smith, Derek J. ; Den Uyl, Paul A. ; Mohamed, Osama G. ; Yu, Fengan ; Ruberg, Steven A. ; Chaffin, Justin D. ; Goodwin, Kelly D. ; Tripathi, Ashootosh ; Sherman, David H. ; et al ( May 2022 , Applied and Environmental Microbiology)

   Rudi, Knut (Ed.)

   ABSTRACT Cyanobacterial harmful algal blooms (cyanoHABs) degrade freshwater ecosystems globally. Microcystis aeruginosa often dominates cyanoHABs and produces microcystin (MC), a class of hepatotoxins that poses threats to human and animal health. Microcystin toxicity is influenced by distinct structural elements across a diversity of related molecules encoded by variant mcy operons. However, the composition and distribution of mcy operon variants in natural blooms remain poorly understood. Here, we characterized the variant composition of mcy genes in western Lake Erie Microcystis blooms from 2014 and 2018. Sampling was conducted across several spatial and temporal scales, including different bloom phases within 2014, extensive spatial coverage on the same day (2018), and frequent, autonomous sampling over a 2-week period (2018). Mapping of metagenomic and metatranscriptomic sequences to reference sequences revealed three Microcystis mcy genotypes: complete (all genes present [ mcyA–J ]), partial (truncated mcyA , complete mcyBC , and missing mcyD–J ), and absent (no mcy genes). We also detected two different variants of mcyB that may influence the production of microcystin congeners. The relative abundance of these genotypes was correlated with pH and nitrate concentrations. Metatranscriptomic analysis revealed that partial operons were, at times, the most abundant genotype and expressed in situ , suggesting the potential biosynthesis of truncated products. Quantification of genetic divergence between genotypes suggests that the observed strains are the result of preexisting heterogeneity rather than de novo mutation during the sampling period. Overall, our results show that natural Microcystis populations contain several cooccurring mcy genotypes that dynamically shift in abundance spatiotemporally via strain succession and likely influence the observed diversity of the produced congeners. IMPORTANCE Cyanobacteria are responsible for producing microcystins (MCs), a class of potent and structurally diverse toxins, in freshwater systems around the world. While microcystins have been studied for over 50 years, the diversity of their chemical forms and how this variation is encoded at the genetic level remain poorly understood, especially within natural populations of cyanobacterial harmful algal blooms (cyanoHABs). Here, we leverage community DNA and RNA sequences to track shifts in mcy genes responsible for producing microcystin, uncovering the relative abundance, expression, and variation of these genes. We studied this phenomenon in western Lake Erie, which suffers annually from cyanoHAB events, with impacts on drinking water, recreation, tourism, and commercial fishing. 

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2. Dissolved nitrogen form mediates phycocyanin content in cyanobacteria

   https://doi.org/10.1111/fwb.13892  

   Gladfelter, Matthew F. ; Buley, Riley P. ; Belfiore, Angelea P. ; Fernandez‐Figueroa, Edna G. ; Gerovac, Bridget L. ; Baker, Nicole D. ; Wilson, Alan E. ( March 2022 , Freshwater Biology)

   Chemically reduced nitrogen forms are increasing in aquatic systems and beginning to reach concentrations not previously measured. Despite this, little research has examined the potential of reduced nitrogen forms to encourage excess nitrogen storage and promote algal bloom longevity compared to oxidised forms.

   A 2‐week field, pulse‐application experiment was carried out using 1,100‐L plastic limnocorrals to examine cyanobacterial community response to three nitrogen forms, including nitrate, ammonium, and urea (added as 600 µg N/L). Cell pigments and counts were used to calculate cell‐specific pigment concentrations, and cell‐associated microcystin concentrations were also measured to examine toxin response to a shift in nitrogen source.

   Results showed that, upon nitrogen introduction, extracellular nitrogen quickly decreased in accordance with an increase in cellular phycocyanin 72 hr after fertilisation. Ammonium and urea treatments had more phycocyanin/cell than nitrate or control treatments at 72 hr. After 72 hr, phycocyanin content quickly decreased, consistent with the use of nitrogen from phycobiliproteins. Despite the decrease in light‐harvesting pigments, the total number of cyanobacterial cells increased in the ammonium and urea treatments after 2 weeks. Cyanobacterial particulate toxin (microcystin) quotas were not affected by nitrogen additions.

   Results show that reduced nitrogen forms encourage greater nitrogen storage as pigments and increase bloom longevity compared to oxidised forms.

   Findings support previous studies that suggest reduced nitrogen forms encourage greater cell density and algal bloom persistence. Results further point to excess nitrogen storage as another mechanism that allows cyanobacteria to dominate freshwater systems despite variable environmental conditions.

    

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3. Episodic Decrease in Temperature Increases mcy Gene Transcription and Cellular Microcystin in Continuous Cultures of Microcystis aeruginosa PCC 7806

   https://doi.org/10.3389/fmicb.2020.601864  

   Martin, Robbie M. ; Moniruzzaman, Mohammad ; Stark, Gwendolyn F. ; Gann, Eric R. ; Derminio, Dominique S. ; Wei, Bofan ; Hellweger, Ferdi L. ; Pinto, Ameet ; Boyer, Gregory L. ; Wilhelm, Steven W. ( December 2020 , Frontiers in Microbiology)

   null (Ed.)

   Microcystins produced during harmful cyanobacterial blooms are a public health concern. Although patterns are emerging, the environmental cues that stimulate production of microcystin remain confusing, hindering our ability to predict fluctuations in bloom toxicity. In earlier work, growth at cool temperatures relative to optimum (18°C vs. 26°C) was confirmed to increase microcystin quota in batch cultures of Microcystis aeruginosa NIES-843. Here, we tested this response in M. aeruginosa PCC 7806 using continuous cultures to examine temporal dynamics and using RNA-sequencing to investigate the physiological nature of the response. A temperature reduction from 26 to 19°C increased microcystin quota ∼2-fold, from an average of ∼464 ag μm –3 cell volume to ∼891 ag μm –3 over a 7–9 d period. Reverting the temperature to 26°C returned the cellular microcystin quota to ∼489 ag μm –3 . Long periods (31–42 d) at 19°C did not increase or decrease microcystin quota beyond that observed at 7–9 d. Nitrogen concentration had little effect on the overall response. RNA sequencing indicated that the decrease in temperature to 19°C induced a classic cold-stress response in M. aeruginosa PCC 7806, but this operated on a different timescale than the increased microcystin production. Microcystin quota showed a strong 48- to 72-h time-lag correlation to mcy gene expression, but no correlation to concurrent mcy expression. This work confirms an effect of temperature on microcystin quota and extends our understanding of the physiological nature of the response. 

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4. Factors influencing urea use by giant kelp ( Macrocystis pyrifera , Phaeophyceae)

   https://doi.org/10.1002/lno.11674  

   Smith, Jason M. ; Blasco, Gordon ; Brzezinski, Mark A. ; Melack, John M. ; Reed, Daniel C. ; Miller, Robert J. ( December 2020 , Limnology and Oceanography)

   Urea is an available and readily used source of nitrogen for giant kelp,Macrocystis pyrifera, but little is known about its potential importance for sustaining growth. Results of kinetic experiments indicate urea uptake saturates at an average maximum rate (V_max) of 0.73–0.92 μmol N g dw⁻¹h⁻¹with a half saturation constant (K_s) of 1.02–1.08 μM. The affinity of giant kelp for urea was high relative to that reported for other seaweeds. However, results of similar kinetics experiments with natural, co‐occurring phytoplankton communities indicate that the rate of urea uptake by phytoplankton was > 10‐fold higher than that of giant kelp. Urea uptake by giant kelp decreased 3–12% in darkness (relative to in light) compared to a 66–85% decline for phytoplankton. Similar differences were observed for ammonium and nitrate, suggesting that light intensity and photocycles influence the outcome of competition for N between giant kelp and phytoplankton. Monthly measures of urease in kelp tissues revealed persistent activity at levels that were 100‐fold higher than rates of urea uptake (0.13–0.35 μmol N g fw⁻¹min⁻¹). This finding, coupled with unsuccessful efforts to induce additional urease activity through substrate additions, suggests that urease plays a role in giant kelp physiology beyond that of processing urea taken up from the environment. Collectively, our results suggest giant kelp uses multiple forms of N including urea to sustain year‐round growth. Its consistent capacity to acquire N during both day and night may help offset its low uptake rates relative to phytoplankton and increase its ability to compete for N during periods of low N availability.

    

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5. Feedback Regulation between Aquatic Microorganisms and the Bloom-Forming Cyanobacterium Microcystis aeruginosa

   https://doi.org/10.1128/AEM.01362-19  

   Zhang, Meng ; Lu, Tao ; Paerl, Hans W. ; Chen, Yiling ; Zhang, Zhenyan ; Zhou, Zhigao ; Qian, Haifeng ; Drake, Harold L. ( November 2019 , Applied and Environmental Microbiology)

   ABSTRACT The frequency and intensity of cyanobacterial blooms are increasing worldwide. Interactions between toxic cyanobacteria and aquatic microorganisms need to be critically evaluated to understand microbial drivers and modulators of the blooms. In this study, we applied 16S/18S rRNA gene sequencing and metabolomics analyses to measure the microbial community composition and metabolic responses of the cyanobacterium Microcystis aeruginosa in a coculture system receiving dissolved inorganic nitrogen and phosphorus (DIP) close to representative concentrations in Lake Taihu, China. M. aeruginosa secreted alkaline phosphatase using a DIP source produced by moribund and decaying microorganisms when the P source was insufficient. During this process, M. aeruginosa accumulated several intermediates in energy metabolism pathways to provide energy for sustained high growth rates and increased intracellular sugars to enhance its competitive capacity and ability to defend itself against microbial attack. It also produced a variety of toxic substances, including microcystins, to inhibit metabolite formation via energy metabolism pathways of aquatic microorganisms, leading to a negative effect on bacterial and eukaryotic microbial richness and diversity. Overall, compared with the monoculture system, the growth of M. aeruginosa was accelerated in coculture, while the growth of some cooccurring microorganisms was inhibited, with the diversity and richness of eukaryotic microorganisms being more negatively impacted than those of prokaryotic microorganisms. These findings provide valuable information for clarifying how M. aeruginosa can potentially modulate its associations with other microorganisms, with ramifications for its dominance in aquatic ecosystems. IMPORTANCE We measured the microbial community composition and metabolic responses of Microcystis aeruginosa in a microcosm coculture system receiving dissolved inorganic nitrogen and phosphorus (DIP) close to the average concentrations in Lake Taihu. In the coculture system, DIP is depleted and the growth and production of aquatic microorganisms can be stressed by a lack of DIP availability. M. aeruginosa could accelerate its growth via interactions with specific cooccurring microorganisms and the accumulation of several intermediates in energy metabolism-related pathways. Furthermore, M. aeruginosa can decrease the carbohydrate metabolism of cooccurring aquatic microorganisms and thus disrupt microbial activities in the coculture. This also had a negative effect on bacterial and eukaryotic microbial richness and diversity. Microcystin was capable of decreasing the biomass of total phytoplankton in aquatic microcosms. Overall, compared to the monoculture, the growth of total aquatic microorganisms is inhibited, with the diversity and richness of eukaryotic microorganisms being more negatively impacted than those of prokaryotic microorganisms. The only exception is M. aeruginosa in the coculture system, whose growth was accelerated. 

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