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Microcystis spp. threaten freshwater ecosystems through the proliferation of cyanobacterial harmful algal blooms (cyanoHABs) and production of the hepatotoxin, microcystin. While microcystin and its biosynthesis pathway, encoded by the mcy genes, have been well studied for over 50 years, a recent study found that Microcystis populations in western Lake Erie contain a transcriptionally active partial mcy operon, in which the A2 domain of mcyA and mcyB-C are present but the mcyD-J genes are absent. Here, we investigate the potential biosynthetic products and the evolutionary history of this partial operon. Our results reveal two candidate tetrapeptide constructs, with an X variable position, to be produced by strains with the partial operon. The partial operon appears necessary and sufficient for tetrapeptide biosynthesis and likely evolved from a single ancestor hundreds to tens of thousands of years ago. Bioactivity screens using Hep3B cells indicate a mild elevation of some markers of hepatotoxicity and inflammation, suggesting the need to further assess the effects of these novel secondary metabolites on freshwater ecosystems and public health. The need to assess these effects is even more pressing given the detection of tetrapeptides in both culture and western Lake Erie, which is a vital source of fresh water. Results from this study emphasize previous findings in which novel bacterial secondary metabolites may be derived from the molecular evolution of existing biosynthetic machinery under different environmental forcings.

Microcystis and bacteria always live together in the mucilage of Microcystis colonies. Extracellular electrons between Microcystis and bacteria can be translated from bioenergy to electric energy. Here, photosynthetic microbial fuel cells (PMFCs) were constructed to make clear the electron transfer mechanism between Microcystis and bacteria. A remarkable enhancement of current density with 2.5-fold change was detected in the coculture of Microcystis and bacteria than pure culture of Microcystis. Transcriptome analyses showed that photosynthesis efficiency of Microcystis was upregulated and may release more electron to improve extracellular electron transfer rate. Significant increase on oxidative phosphorylation of bacterial community was observed according to meta-transcriptome. Bacterial electrons were transferred out of cell membranes by enhancing VgrG and IcmF copies though the type II bacterial secretion system. Not only Microcystis and bacteria attached with each other tightly by filamentous, but also more gene copies relating to pilin and riboflavin production were detected from Microcystis culture. A confirmatory experiment found that riboflavin can upregulate the electron transfer and current density by adding riboflavin into cocultures. Thus, the direct contact and indirect interspecies electron transfer processes between Microcystis and bacteria were observed. Results enlarge knowledge for activities of Microcystis colonies in cyanobacterial blooms, and provide a better understanding for energy transformation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

As a key form of post-transcriptional regulation, microRNAs (miRNAs) regulate gene expression by binding to target mRNAs, leading to mRNA decay or translational repression. Recently, the role of miRNAs in the response of aquatic organisms to environmental stressors has emerged. Daphnia , widely distributed cladocerans, play a crucial role in aquatic ecosystems. Cyanobacterial blooms often cause Daphnia populations to decrease, thereby disrupting ecosystem functionality and water quality. However, the post-transcriptional mechanisms behind Daphnia 's response to toxic cyanobacteria are insufficiently understood. This study investigated the role of miR-210, a multifunctional miRNA involved in stress response and toxicity pathways, and its target genes ( MLH3 , CDHR5 , and HYOU1 ) in two Daphnia magna clones exposed to toxic Microcystis aeruginosa . Results showed that M. aeruginosa inhibited somatic growth rates, led to microcystin accumulation, caused abnormal ultrastructural alterations in the digestive tract, and induced DNA damage in both clones. Notably, exposure significantly increased miR-210 expression and decreased the expression of its target genes compared with the controls. We identified miR-210s regulation on clonal-tolerance variations in D. magna to M. aeruginosa , emphasizing miRNAs' contribution to adaptive responses. Our work uncovered a novel post-transcriptional mechanism of cyanobacterial impact on zooplankton and provided essential insights for assessing cyanobacterial toxicity risks.

Cyanobacterial blooms are expanding world-wide in freshwater and marine environments, and can cause serious ecological and environmental issues, which also contribute to the spread of antibiotic resistance genes (ARGs). However, the mechanistic understanding of cyanobacteria-mediated resistance dynamics is not fully elucidated yet. We selected Microcystis aeruginosa as a model cyanobacteria to illustrate how cyanobacteria mediate the evolution and transfer processes of bacterial antibiotic resistance. The results show that the presence of cyanobacteria significantly decreased the abundance of antibiotic resistant bacteria (ARB) and antibiotic resistant genes (ARGs) by 3%-99% and 2%-18%, respectively. In addition, it clearly altered bacterial community structure, with the dominant genera evolving from Acinetobacter (27%) and Enterobacter (42%) to Porphyrobacter (59%). The abundance of ARGs positively correlated with Proteobacteria and Firmicutes, rather than Cyanobacteria, and Bacteroidetes. In the presence of cyanobacteria, the transfer events of bacterial resistance genes via conjugation were found to decrease by 10%-89% (p < 0.05). Surprisingly, we found an extradentary high transfer frequency (about 0.1) for the ARGs via plasmid conjugation from the bacteria into M. aeruginosa population. It confirmed the role of cyanobacterial population as the competent hosts to facilitate ARGs spreading. Our findings provide valuable information on the risk evaluation of ARGs caused by cyanobacterial blooms in aquatic environments, key for the protection and assessment of aquatic environmental quality., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The Microcystis mobilome is a well-known but understudied component of this bloom-forming cyanobacterium. Through genomic and transcriptomic comparisons, we found five families of transposases that altered the expression of genes in the well-studied toxigenic type-strain, Microcystis aeruginosa PCC 7086, and a non-toxigenic genetic mutant, Microcystis aeruginosa PCC 7806 ΔmcyB. Since its creation in 1997, the ΔmcyB strain has been used in comparative physiology studies against the wildtype strain by research labs throughout the world. Some differences in gene expression between what were thought to be otherwise genetically identical strains have appeared due to insertion events in both intra- and intergenic regions. In our ΔmcyB isolate, a sulfate transporter gene cluster (sbp-cysTWA) showed differential expression from the wildtype, which may have been caused by the insertion of a miniature inverted repeat transposable element (MITE) in the sulfate-binding protein gene (sbp). Differences in growth in sulfate-limited media also were also observed between the two isolates. This paper highlights how Microcystis strains continue to "evolve" in lab conditions and illustrates the importance of insertion sequences / transposable elements in shaping genomic and physiological differences between Microcystis strains thought otherwise identical. This study forces the necessity of knowing the complete genetic background of isolates in comparative physiological experiments, to facilitate the correct conclusions (and caveats) from experiments., (© 2024. The Author(s).)

We aimed to evaluate the ability of naturally occurring colonies of Microcystis, embedded in a thick mucilage, to persist in estuarine waters. In two batch experiments, we examined the dynamics of microbial communities, including cyanobacteria and associated heterotrophic bacteria, sampled from the field during both a cyanobacterial bloom (non-limiting nutrient condition) and the post-bloom period (limiting nutrient condition), and subjected them to a salinity gradient representative of the freshwater-marine continuum. We demonstrated that both Microcystis aeruginosa and M. wesenbergii survived high salinities due to osmolyte accumulation. Specifically, prolonged exposure to high salinity led to betaine accumulation in the cyanobacterial biomass. The relative abundance of the mcyB gene remained around 30%, suggesting no selection for toxic genotypes with salinity or nutrient changes. Microcystins were predominantly intracellular, except at high salinity levels (>15), where more than 50% of the total microcystin concentration was extracellular. In both nutrient conditions, over 70% of the heterotrophic bacterial community belonged to the Gammaproteobacteria family, followed by the Bacteroidota. Bacterial community composition differed in both size fractions, as well as along the salinity gradient over time. Finally, genus-specific core microbiomes were identified and conserved even under highly stressful conditions, suggesting interactions that support community stability and resilience., (© 2024 The Author(s). Environmental Microbiology Reports published by John Wiley & Sons Ltd.)

The local ecosystems, fishery and human health are all threatened by water blooms, so effectively controlling water blooms has become an urgent and challenging issue. Biological control of water blooms is given priority due to its low cost, high efficiency and environmental friendliness. In this study, Pseudomonas ZY-1 and Bacillus FY-1, two highly-effective algicidal bacteria strains which are able to indirectly lyse algae by separating and screening from the vigorous water body in the paddy alga of Northeast China were obtained. The two bacterial strains have stronger ability to lyse alga in the bacterial liquid concentration of 10 6 CFU/ml, and the alga-lysing rate on 7 d reached 84.03% and 83.11% respectively. The active substance secreted by ZY-1 is not sensitive to the changes of temperature and pH value, while as FY-1 cell-free filtrate is not stable in high temperature above 50 ℃ and pH of 5, it requires the sun light to have the algaecidal effect. The cell-free filtrates of strains ZY-1 and FY-1 had the best lysis effect on Microcystis aeruginosa cells, and the chlorophyll a content of algae decreased to 0.13 ± 0.02 mg/L and 0.14 ± 0.03 mg/L respectively and the Fv/Fm of Microcystis aeruginosa decreased by 97.22% after 7 days. The algaecidal process of ZY-1 and FY-1 may be that the cell-free filtrate inhibits the photosynthesis of Microcystis aeruginosa, and meanwhile it avoids the regeneration and repair of photosynthesis of algal cells by affecting the gene expression and damaging the repair system of algal cells, so the membrane lipid peroxidation is exacerbated and then the membrane of algal cells is broken, the algal cells can't do normal life activities, and finally the algal cell would be killed. The rice seedlings in the algal liquid treatment group are short and show root dysplasia, few roots and brown roots. After treated with cell-free filtrate of ZY-1 and FY-1, the oxidative damage of the rice is obviously reduced, and the harm from Microcystis aeruginosa is reduced, which has the repair effect to the roots of rice seedlings and its aboveground growth. The cell-free filtrate of FY-1 works better than ZY-1. The bacteria strains of ZY-1 and FY-1 have the indirect algaecide trait, which makes them the potential environmentally-friendly algaecidal bacteria and they show broad application in the agricultural production and the control of water blooms., (© 2024. The Author(s).)

The rapid growth of cyanobacteria, particularly Microcystis aeruginosa, poses a significant threat to global water security. The proliferation of toxic Microcystis aeruginosa raises concerns due to its potential harm to human health and socioeconomic impacts. Dense blooms contribute to spatiotemporal inorganic carbon depletion, promoting interest in the roles of carbon-concentrating mechanisms (CCMs) for competitive carbon uptake. Despite the importance of HCO 3 - transporters, genetic evaluations and functional predictions in M. aeruginosa remain insufficient. In this study, we explored the diversity of HCO 3 - transporters in the genomes of 46 strains of M. aeruginosa, assessing positive selection for each. Intriguingly, although the Microcystis BicA transporter became a partial gene in 23 out of 46 genomic strains, we observed significant positive sites. Structural analyses, including predicted 2D and 3D models, confirmed the structural conservation of the Microcystis BicA transporter. Our findings suggest that the Microcystis BicA transport likely plays a crucial role in competitive carbon uptake, emphasizing its ecological significance. The ecological function of the Microcystis BicA transport in competitive growth during cyanobacterial blooms raises important questions. Future studies require experimental confirmation to better understand the role of the Microcysits BicA transporter in cyanobacterial blooms dynamics., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Despite significant reductions in phosphorus (P) loads, lakes still experience cyanobacterial blooms. Little is known regarding cellular P regulation in response to P deficiency in widely distributed bloom causing species such as Microcystis. In this study, we investigated changes in P containing and non-P lipids contents and their ratios concomitantly with the determinations of expression levels of genes encoding these lipids in cultural and field Microcystis samples. In the culture, the content of phosphatidylglycerol (PG) decreased from 2.1 μg g -1 in P replete control to 1.2 μg g -1 in P-deficient treatment, while non-P lipids, like sulfoquinovosyldiacylglycerol (SQDG) and monogalactosyldiacylglycerol (MGDG), increased dramatically from 13.6 μg g -1 to 142.3 μg g -1 , and from 0.9 μg g -1 to 16.74 μg g -1 , respectively. The expression of the MGDG synthesis gene, mgdE, also increased under low P conditions. Significant positive relationships between soluble reactive phosphorus (SRP) and ratios of P-containing lipids (PG) to non-P lipids, including SQDG, MGDG and digalactosyldiacylglycerol (DGDG) (P < 0.05) were observed in the field investigations. Both cultural and field data indicated that Microcystis sp. might increase non-P lipids proportion to lower P demand when suffering from P deficiency. Furthermore, despite lipid remodeling, photosynthetic activity remained stable, as indicated by comparable chlorophyll fluorescence and Fv/Fm ratios among cultural treatments. These findings suggested that Microcystis sp. may dominate in P-limited environments by substituting glycolipids and sulfolipids for phospholipids to reduce P demand without compromising the photosynthetic activity. This effective strategy in response to P deficiency meant a stricter P reduction threshold is needed in terms of Microcystis bloom control., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

The toxicity of triclosan (TCS) to various aquatic organisms has been demonstrated at environmental concentrations. However, the effects and mechanisms of TCS on toxic cyanobacteria remains largely unexplored. This study investigated the physiological and molecular variations in two representative toxic Microcystis species (M. aeruginosa and M. viridis) under exposure to TCS for 12 d. Our findings demonstrated that the median effective concentration (EC 50 ) of TCS for both Microcystis species were close to the levels detected in the environment (M. aeruginosa: 9.62 μg L -1 ; M. viridis: 27.56 μg L -1 ). An increased level of reactive oxygen species (ROS) was observed in Microcystis, resulting in oxidative damage when exposed to TCS at concentrations ranging from 10 μg L -1 to 50 μg L -1 . The photosynthetic activity of Microcystis had a certain degree of recovery capability at low concentrations of TCS. Compared to M. aeruginosa, the higher recovery capability of the photosynthetic system in M. viridis would be mainly attributed to the increased ability for PSII repair and phycobilisome synthesis. Additionally, the synthesis of microcystins in the two species and the release rate in M. viridis significantly increased under 10-50 μg L -1 TCS. At the molecular level, exposure to TCS at EC 50 for 12 d induced the dysregulation of genes associated with photosynthesis and antioxidant system. The upregulation of genes associated with microcystin synthesis and nitrogen metabolism further increased the potential risk of microcystin release. Our results revealed the aquatic toxicity and secondary ecological risks of TCS at environmental concentrations, and provided theoretical data with practical reference value for TCS monitoring., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The massive proliferation of Microcystis threatens freshwater ecosystems and degrades water quality globally. Understanding the mechanisms that contribute to Microcystis growth is crucial for managing Microcystis blooms. The lifestyles of bacteria can be classified generally into two groups: particle-attached (PA; > 3 µm) and free-living (FL; 0.2-3.0 µm). However, little is known about the response of PA and FL bacteria to Microcystis blooms. Using 16S rRNA gene high-throughput sequencing, we investigated the stability, assembly process, and co-occurrence patterns of PA and FL bacterial communities during distinct bloom stages. PA bacteria were phylogenetically different from their FL counterparts. Microcystis blooms substantially influenced bacterial communities. The time decay relationship model revealed that Microcystis blooms might increase the stability of both PA and FL bacterial communities. A contrasting community assembly mechanism was observed between the PA and FL bacterial communities. Throughout Microcystis blooms, homogeneous selection was the major assembly process that impacted the PA bacterial community, whereas drift explained much of the turnover of the FL bacterial community. Both PA and FL bacterial communities could be separated into modules related to different phases of Microcystis blooms. Microcystis blooms altered the assembly process of PA and FL bacterial communities. PA bacterial community appeared to be more responsive to Microcystis blooms than FL bacteria. Decomposition of Microcystis blooms may enhance cooperation among bacteria. Our findings highlight the importance of studying bacterial lifestyles to understand their functions in regulating Microcystis blooms. KEY POINTS: • Microcystis blooms alter the assembly process of PA and FL bacterial communities • Microcystis blooms increase the stability of both PA and FL bacterial communities • PA bacteria seem to be more responsive to Microcystis blooms than FL bacteria., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Microcystis, a type of cyanobacteria known for producing microcystins (MCs), is experiencing a global increase in blooms. They have been recently recognized as potential contributors to the widespread of antibiotic resistance genes (ARGs). By reviewing approximately 150 pieces of recent studies, a hypothesis has been formulated suggesting that significant fluctuations in MCs concentrations and microbial community structure during Microcystis blooms could influence the dynamics of waterborne ARGs. Among all MCs, microcystin-LR (MC-LR) is the most widely distributed worldwide, notably abundant in reservoirs during summer. MCs inhibit protein phosphatases or increase reactive oxygen species (ROS), inducing oxidative stresses, enhancing membrane permeability, and causing DNA damage. This further enhances selective pressures and horizontal gene transfer (HGT) chances of ARGs. The mechanisms by which Microcystis regulates ARG dissemination have been systematically organized for the first time, focusing on the secretion of MCs and the alterations of bacterial community structure. However, several knowledge gaps remain, particularly concerning how MCs interfere with the electron transport chain and how Microcystis facilitates HGT of ARGs. Concurrently, the predominance of Microcystis forming the algal microbial aggregates is considered a hotspot for preserving and transferring ARGs. Yet, Microcystis can deplete the nutrients from other taxa within these aggregates, thereby reducing the density of ARG-carrying bacteria. Therefore, further studies are needed to explore the 'symbiotic - competitive' relationships between Microcystis and ARG-hosting bacteria under varied nutrient conditions. Addressing these knowledge gaps is crucial to understand the impacts of the algal aggregates on dynamics of waterborne antibiotic resistome, and underscores the need for effective control of Microcystis to curb the spread of antibiotic resistance. Constructed wetlands and photocatalysis represent advantageous strategies for halting the spread of ARGs from the perspective of Microcystis blooms, as they can effectively control Microcystis and MCs while maintaining the stability of aquatic ecosystem., Competing Interests: Declaration of competing interest Authors read and approved the final manuscript. There are no competing interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Biological networks serve a crucial role in elucidating intricate biological processes. While interspecies environmental interactions have been extensively studied, the exploration of gene interactions within species, particularly among individual microorganisms, is less developed. The increasing amount of microbiome genomic data necessitates a more nuanced analysis of microbial genome structures and functions. In this context, we introduce a complex structure using higher-order network theory, "Solid Motif Structures (SMS)", via a hierarchical biological network analysis of genomes within the same genus, effectively linking microbial genome structure with its function. Leveraging 162 high-quality genomes of Microcystis, a key freshwater cyanobacterium within microbial ecosystems, we established a genome structure network. Employing deep learning techniques, such as adaptive graph encoder, we uncovered 27 critical functional subnetworks and their associated SMSs. Incorporating metagenomic data from seven geographically distinct lakes, we conducted an investigation into Microcystis' functional stability under varying environmental conditions, unveiling unique functional interaction models for each lake. Our work compiles these insights into an extensive resource repository, providing novel perspectives on the functional dynamics within Microcystis. This research offers a hierarchical network analysis framework for understanding interactions between microbial genome structures and functions within the same genus., (© 2024. The Author(s).)

High levels of environmental H 2 O 2 represent a threat to many freshwater bacterial species, including toxic-bloom-forming Microcystis aeruginosa, particularly under high-intensity light conditions. The highest extracellular catalase activity-possessing Pseudoduganella aquatica HC52 was chosen among 36 culturable symbiotic isolates from the phycosphere in freshly collected M. aeruginosa cells. A zymogram for catalase activity revealed the presence of only one extracellular catalase despite the four putative catalase genes (katA1, katA2, katE, and srpA) identified in the newly sequenced genome (∼6.8 Mb) of P. aquatica HC52. Analysis of secreted catalase using liquid chromatography-tandem mass spectrometry was identified as KatA1, which lacks a typical signal peptide, although the underlying mechanism for its secretion is unknown. The expression of secreted KatA1 appeared to be induced in the presence of H 2 O 2 . Proteomic analysis also confirmed the presence of KatA1 inside the outer membrane vesicles secreted by P. aquatica HC52 following exposure to H 2 O 2 . High light intensities (> 100 µmol m -2 s -1 ) are known to kill catalase-less axenic M. aeruginosa cells, but the present study found that the presence of P. aquatica cells supported the growth of M. aeruginosa, while the extracellular catalases in supernatant or purified form also sustained the growth of M. aeruginosa under the same conditions. Our results suggest that the extracellular catalase secreted by P. aquatica HC52 enhances the tolerance of M. aeruginosa to H 2 O 2 , thus promoting the formation of M. aeruginosa blooms under high light intensities., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Microcystis spp. are renowned for producing the hepatotoxin microcystin in freshwater cyanobacterial harmful algal blooms around the world, threatening drinking water supplies and public and environmental health. However, Microcystis genomes also harbor numerous biosynthetic gene clusters (BGCs) encoding the biosynthesis of other secondary metabolites, including many with toxic properties. Most of these BGCs are uncharacterized and currently lack links to biosynthesis products. However, recent field studies show that many of these BGCs are abundant and transcriptionally active in natural communities, suggesting potentially important yet unknown roles in bloom ecology and water quality. Here, we analyzed 21 xenic Microcystis cultures isolated from western Lake Erie to investigate the diversity of the biosynthetic potential of this genus. Through metabologenomic and in silico approaches, we show that these Microcystis strains contain variable BGCs, previously observed in natural populations, and encode distinct metabolomes across cultures. Additionally, we find that the majority of metabolites and gene clusters are uncharacterized, highlighting our limited understanding of the chemical repertoire of Microcystis spp. Due to the complex metabolomes observed in culture, which contain a wealth of diverse congeners as well as unknown metabolites, these results underscore the need to deeply explore and identify secondary metabolites produced by Microcystis beyond microcystins to assess their impacts on human and environmental health.IMPORTANCEThe genus Microcystis forms dense cyanobacterial harmful algal blooms (cyanoHABs) and can produce the toxin microcystin, which has been responsible for drinking water crises around the world. While microcystins are of great concern, Microcystis also produces an abundance of other secondary metabolites that may be of interest due to their potential for toxicity, ecological importance, or pharmaceutical applications. In this study, we combine genomic and metabolomic approaches to study the genes responsible for the biosynthesis of secondary metabolites as well as the chemical diversity of produced metabolites in Microcystis strains from the Western Lake Erie Culture Collection. This unique collection comprises Microcystis strains that were directly isolated from western Lake Erie, which experiences substantial cyanoHAB events annually and has had negative impacts on drinking water, tourism, and industry., Competing Interests: C.E.Y. was a graduate student at the time this research was completed but is now an employee at New England Biolabs, Ipswich, MA, USA.

Large and temperate Lake Peipsi is the fourth largest lake in Europe, where the massive cyanobacterial blooms are composed mostly of Microcystis spp., which have been common for several decades now. The seasonal dynamics of potentially toxic Microcystis were studied using microscopy and quantitative polymerase chain reaction (qPCR) by assessing the microcystin-encoding microcystin synthetase gene E (mcyE) abundances. Water samples were analyzed over the lake areas, varying in depth, trophic level, and cyanobacterial composition during the growing period of 2021. The Microcystis mcyE genes were detected through the growing period (May-October), forming peak abundances in September with decreasing temperatures (8.9-11.1 °C). Total phosphorus (TP) and nitrate (NO 3 - ) were the most relevant environmental variables influencing the Microcystis biomass as well as mcyE abundances. Comparison with previous years (2011, 2012) indicated that the abundance and seasonal dynamics of toxigenic Microcystis can be highly variable between the years and lake areas, varying also in dominant Microcystis species. Contrary to expectations, based on mcyE abundances, the increased risk of toxin-producing Microcystis can occur in Peipsi through the growing period, independently of the water temperature and biomasses of Microcystis., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

In this study, the toxigenic characteristics of 14 strains of Microcystis were analyzed, and single nucleotide polymorphism (SNP) and insertion/deletion (InDel) loci in microcystin synthetase (mcy) gene clusters were screened. Based on SNP and InDel loci associated with the toxigenic characteristics, primers and TaqMan or Cycling fluorescent probes were designed to develop duplex real-time fluorescent quantitative PCR (FQ-PCR) assays. After evaluating specificity and sensitivity, these assays were applied to detect the toxigenic Microcystis genotypes in a shrimp pond where Microcystis blooms occurred. The results showed a total of 2155 SNP loci and 66 InDel loci were obtained, of which 12 SNP loci and 5 InDel loci were associated with the toxigenic characteristics. Three duplex real-time FQ-PCR assays were developed, each of which could quantify two genotypes of toxigenic Microcystis. These FQ-PCR assays were highly specific, and two Cycling assays were more sensitive than TaqMan assay. In the shrimp pond, six genotypes of toxigenic Microcystis were detected using the developed FQ-PCR assays, indicating that above genotyping assays have the potential for quantitative analysis of the toxigenic Microcystis genotypes in natural water., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Increasing antivirals in surface water caused by their excessive consumption pose serious threats to aquatic organisms. Our recent research found that the input of antiviral drug arbidol to algal bloom water can induce acute toxicity to the growth and metabolism of Microcystis aeruginosa, resulting in growth inhibition, as well as decrease in chlorophyll and ATP contents. However, the toxic mechanisms involved remained obscure, which were further investigated through transcriptomic analysis in this study. The results indicated that 885-1248 genes in algae were differentially expressed after exposure to 0.01-10.0 mg/L of arbidol, with the majority being down-regulated. Analysis of commonly down-regulated genes found that the cellular response to oxidative stress and damaged DNA bonding were affected, implying that the stress defense system and DNA repair function of algae might be damaged. The down-regulation of genes in porphyrin metabolism, photosynthesis, carbon fixation, glycolysis, tricarboxylic acid cycle, and oxidative phosphorylation might inhibit chlorophyll synthesis, photosynthesis, and ATP supply, thereby hindering the growth and metabolism of algae. Moreover, the down-regulation of genes related to nucleotide metabolism and DNA replication might influence the reproduction of algae. These findings provided effective strategies to elucidate toxic mechanisms of contaminants on algae in algal bloom water., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Aims: Developing energy-saving and ecofriendly strategies for treating harvested Microcystis biomass., Methods and Results: Streptomyces amritsarensis HG-16 was first reported to effectively kill various morphotypes of natural Microcystis colonies at very high cell densities. Concurrently, HG-16 grown on lysed Microcystis maintained its antagonistic activity against plant pathogenic fungus Fusarium graminearum. It could completely inhibit spore germination and destroy mycelial structure of F. graminearum. Transcriptomic analysis revealed that HG-16 attacked F. graminearum in a comprehensive way: interfering with replication, transcription, and translation processes, inhibiting primary metabolisms, hindering energy production and simultaneously destroying stress-resistant systems of F. graminearum., Conclusions: The findings of this study provide a sustainable and economical option for resource reclamation from Microcystis biomass: utilizing Microcystis slurry to propagate HG-16, which can subsequently be employed as a biocontrol agent for managing F. graminearum., (© The Author(s) 2024. Published by Oxford University Press on behalf of Applied Microbiology International.)

The cyanosiphophage Mic1 specifically infects the bloom-forming Microcystis aeruginosa FACHB 1339 from Lake Chaohu, China. Previous genomic analysis showed that its 92,627 bp double-stranded DNA genome consists of 98 putative open reading frames, 63% of which are of unknown function. Here, we investigated the transcriptome dynamics of Mic1 and its host using RNA sequencing. In the early, middle, and late phases of the 10 h lytic cycle, the Mic1 genes are sequentially expressed and could be further temporally grouped into two distinct clusters in each phase. Notably, six early genes, including gp49 that encodes a TnpB-like transposase, immediately reach the highest transcriptional level in half an hour, representing a pioneer cluster that rapidly regulates and redirects host metabolism toward the phage. An in-depth analysis of the host transcriptomic profile in response to Mic1 infection revealed significant upregulation of a polyketide synthase pathway and a type III-B CRISPR system, accompanied by moderate downregulation of the photosynthesis and key metabolism pathways. The constant increase of phage transcripts and relatively low replacement rate over the host transcripts indicated that Mic1 utilizes a unique strategy to gradually take over a small portion of host metabolism pathways after infection. In addition, genomic analysis of a less-infective Mic1 and a Mic1-resistant host strain further confirmed their dynamic interplay and coevolution via the frequent horizontal gene transfer. These findings provide insights into the mutual benefit and symbiosis of the highly polymorphic cyanobacteria M. aeruginosa and cyanophages., Importance: The highly polymorphic Microcystis aeruginosa is one of the predominant bloom-forming cyanobacteria in eutrophic freshwater bodies and is infected by diverse and abundant cyanophages. The presence of a large number of defense systems in M. aeruginosa genome suggests a dynamic interplay and coevolution with the cyanophages. In this study, we investigated the temporal gene expression pattern of Mic1 after infection and the corresponding transcriptional responses of its host. Moreover, the identification of a less-infective Mic1 and a Mic1-resistant host strain provided the evolved genes in the phage-host coevolution during the multiple-generation cultivation in the laboratory. Our findings enrich the knowledge on the interplay and coevolution of M. aeruginosa and its cyanophages and lay the foundation for the future application of cyanophage as a potential eco-friendly and bio-safe agent in controlling the succession of harmful cyanobacterial blooms., Competing Interests: The authors declare no conflict of interest.

The occurrence of toxic bloom-forming cyanobacteria, Microcystis aeruginosa, has been frequently reported worldwide. These colony forming toxic cyanobacteria harbour a wide range of heterotrophic bacterial communities. The present study has attempted to understand the bloom dynamics of M. aeruginosa along with isolating their colony-associated culturable heterotrophic bacteria from two freshwater ponds in south India with a persisting cyanobacterial bloom. The monthly monitoring of these study areas revealed the conducive role of warm, stagnant waters with high nutrients in forming M. aeruginosa bloom. The peak values of temperature, nitrate, and phosphate at station 1 reached up to 30.5 °C, 4.48 mg/L, 1.64 mg/L, and at station 2, 31 °C, 3.45 mg/L, and 0.62 mg/L, respectively. Twenty-eight bacterial isolates belonging to Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, and Firmicutes were obtained during the study. Among these 28 isolates, Firmicutes was dominant with the M. aeruginosa bloom from both the study areas., (© 2023. Institute of Microbiology, Academy of Sciences of the Czech Republic, v.v.i.)

Background: During the bloom season, the colonial cyanobacterium Microcystis forms complex aggregates which include a diverse microbiome within an exopolymer matrix. Early research postulated a simple mutualism existing with bacteria benefitting from the rich source of fixed carbon and Microcystis receiving recycled nutrients. Researchers have since hypothesized that Microcystis aggregates represent a community of synergistic and interacting species, an interactome, each with unique metabolic capabilities that are critical to the growth, maintenance, and demise of Microcystis blooms. Research has also shown that aggregate-associated bacteria are taxonomically different from free-living bacteria in the surrounding water. Moreover, research has identified little overlap in functional potential between Microcystis and members of its microbiome, further supporting the interactome concept. However, we still lack verification of general interaction and know little about the taxa and metabolic pathways supporting nutrient and metabolite cycling within Microcystis aggregates., Results: During a 7-month study of bacterial communities comparing free-living and aggregate-associated bacteria in Lake Taihu, China, we found that aerobic anoxygenic phototrophic (AAP) bacteria were significantly more abundant within Microcystis aggregates than in free-living samples, suggesting a possible functional role for AAP bacteria in overall aggregate community function. We then analyzed gene composition in 102 high-quality metagenome-assembled genomes (MAGs) of bloom-microbiome bacteria from 10 lakes spanning four continents, compared with 12 complete Microcystis genomes which revealed that microbiome bacteria and Microcystis possessed complementary biochemical pathways that could serve in C, N, S, and P cycling. Mapping published transcripts from Microcystis blooms onto a comprehensive AAP and non-AAP bacteria MAG database (226 MAGs) indicated that observed high levels of expression of genes involved in nutrient cycling pathways were in AAP bacteria., Conclusions: Our results provide strong corroboration of the hypothesized Microcystis interactome and the first evidence that AAP bacteria may play an important role in nutrient cycling within Microcystis aggregate microbiomes. Video Abstract., (© 2024. The Author(s).)

Hydrogen peroxide is a reactive oxygen species (ROS) naturally occurring at low levels in aquatic environments and production varies widely across different ecosystems. Oxygenic photosynthesis generates hydrogen peroxide as a byproduct, of which some portion can be released to ambient water. However, few studies have examined hydrogen peroxide dynamics in relation to cyanobacterial harmful algal blooms (cHABs). A year-long investigation of algal succession and hydrogen peroxide dynamics was conducted at the Caloosahatchee River, Florida, USA. We aimed to identify potential biological mechanisms responsible for elevated hydrogen peroxide production during cHAB events through the exploration of the freshwater microbial metatranscriptome. Hydrogen peroxide concentrations were elevated from February to September of 2021 when cyanobacteria were active and abundant. We observed one Microcystis cHAB event in spring and one in winter. Both had distinct nutrient uptake and cyanotoxin gene expression patterns. While meaningful levels of microcystin were only detected during periods of elevated hydrogen peroxide, cyanopeptolin was by far the most expressed cyanotoxin during the spring bloom when hydrogen peroxide was at its yearly maxima. Gene expressions of five microbial enzymes (Rubisco, superoxide dismutase, cytochrome b559, pyruvate oxidase, and NADH dehydrogenase) positively correlated to hydrogen peroxide concentrations. Additionally, there was higher nitrogen-fixing gene (nifDKH) expression by filamentous cyanobacteria after the spring bloom but no secondary bloom formation occurred. Overall, elevated environmental hydrogen peroxide concentrations were linked to cyanobacterial dominance and greater expression of specific enzymes in the photosynthesis of cyanobacteria. This implicates cyanobacterial photosynthesis and growth results in increased hydrogen peroxide generation as reflected in measured environmental concentrations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

An alpha-proteobacterial strain JXJ CY 53  T was isolated from the cyanosphere of Microcystis sp. FACHB-905 (MF-905) collected from Lake Dianchi, China. JXJ CY 53  T was observed to be an aerobic, Gram-stain-negative, oval shaped, and mucus-secreting bacterium. It had C 18:1 ω7c and C 16:0 as the major cellular fatty acids, Q-10 as the predominant ubiquinone, and sphingoglycolipid, diphosphatidylglycerol, phosphatidylcholine, and phosphatidylmethylethanolamine as the polar lipids. The G + C content of DNA was 65.85%. The bacterium had 16S rRNA gene sequence identities of 98.9% and 98.7% with Sphingomonas panni DSM 15761  T and Sphingomonas hankookensis KCTC 22579  T , respectively, while less than 97.4% identities with other members of the genus. Further taxonomic analysis indicated that JXJ CY 53  T represented a new member of Sphingomonas, and the species epithet was proposed as Sphingomonas lacusdianchii sp. nov. (type strain JXJ CY 53  T  = KCTC 72813  T  = CGMCC 1.17657  T ). JXJ CY 53  T promoted the growth of MF-905 by providing bio-available phosphorus and nitrogen, plant hormones, vitamins, and carotenoids. It could modulate the relative abundances of nonculturable bacteria associated with MF-905 and influence the interactions of MF-905 and other bacteria isolated from the cyanobacterium, in addition to microcystin production characteristics. Meanwhile, MF-905 could provide JXJ CY 53  T dissolved organic carbon for growth, and control the growth of JXJ CY 53  T by secreting specific chemicals other than microcystins. Overall, these results suggest that the interactions between Microcystis and its attached bacteria are complex and dynamic, and may influence the growth characteristics of the cyanobacterium. This study provided new ideas to understand the interactions between Microcystis and its attached bacteria. KEY POINTS: • A novel bacterium (JXJCY 53 T ) was isolated from the cyanosphere of Microcystis sp. FACHB-905 (MF-905) • JXJCY 53 T modulated the growth and microcystin production of MF-905 • MF-905 could control the attached bacteria by specific chemicals other than microcystins (MCs)., (© 2024. The Author(s).)

Short-chain Perfluorinated compounds (PFCs), used as substitutes for highly toxic long-chain PFCs, are increasingly entering the aquatic environment. However, the toxicity of short-chain PFCs in the environment is still controversial. This study investigated the effects of short-chain perfluorobutanesulfonic acid (PFBS) at different concentrations (2.5, 6, 14.4, 36, and 90 mg/L) on M. aeruginosa growth under 12-day exposure and explored the molecular mechanism of toxicity using transcriptomics. The results showed that M. aeruginosa exhibited hormetic effects after exposure to PFBS. Low PFBS concentrations stimulated algal growth, whereas high PFBS concentrations inhibited it, and this inhibitory effect became progressively more pronounced with increasing PFBS exposure concentrations. Transcriptomics showed that PFBS promoted the pathways of photosynthesis, glycolysis, energy metabolism and peptidoglycan synthesis, providing the energy required for cell growth and maintaining cellular morphology. PFBS, on the other hand, caused growth inhibition in algae mainly through oxidative stress, streptomycin synthesis, and genetic damage. Our findings provide new insights into the toxicity and underlying mechanism of short-chain PFCs on algae and inform the understanding of the hormetic effect of short-chain PFCs, which are crucial for assessing their ecological risks in aquatic environments., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Detecting cyanobacteria in environments is an important concern due to their crucial roles in ecosystems, and they can form blooms with the potential to harm humans and nonhuman entities. However, the most widely used methods for high-throughput detection of environmental cyanobacteria, such as 16S rRNA sequencing, typically provide above-species-level resolution, thereby disregarding intraspecific variation. To address this, we developed a novel DNA microarray tool, termed the CyanoStrainChip, that enables strain-level comprehensive profiling of environmental cyanobacteria. The CyanoStrainChip was designed to target 1277 strains; nearly all major groups of cyanobacteria are included by implementing 43,666 genome-wide, strain-specific probes. It demonstrated strong specificity by in vitro mock community experiments. The high correlation (Pearson's R > 0.97) between probe fluorescence intensities and the corresponding DNA amounts (ranging from 1-100 ng) indicated excellent quantitative capability. Consistent cyanobacterial profiles of field samples were observed by both the CyanoStrainChip and next-generation sequencing methods. Furthermore, CyanoStrainChip analysis of surface water samples in Lake Chaohu uncovered a high intraspecific variation of abundance change within the genus Microcystis between different severity levels of cyanobacterial blooms, highlighting two toxic Microcystis strains that are of critical concern for Lake Chaohu harmful blooms suppression. Overall, these results suggest a potential for CyanoStrainChip as a valuable tool for cyanobacterial ecological research and harmful bloom monitoring to supplement existing techniques.

Species of the Microcystis genus are the most common bloom-forming toxic cyanobacteria worldwide. They belong to a clade of unicellular cyanobacteria whose ability to reach high biomasses during blooms is linked to the formation of colonies. Colonial lifestyle provides several advantages under stressing conditions of light intensity, ultraviolet light, toxic substances and grazing. The progression from a single-celled organism to multicellularity in Microcystis has usually been interpreted as individual phenotypic responses of the cyanobacterial cells to the environment. Here, we synthesize current knowledge about Microcystis colonial lifestyle and its role in the organism ecology. We then briefly review the available information on Microcystis microbiome and propose that changes leading from single cells to colonies are the consequence of specific and tightly regulated signals between the cyanobacterium and its microbiome through a biofilm-like mechanism. The resulting colony is a multi-specific community of interdependent microorganisms., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

Hydrogen peroxide has gained popularity as an environmentally friendly treatment for cyanobacterial harmful algal blooms (cHABs) that takes advantage of oxidative stress sensitivity in cyanobacteria at controlled concentrations. Higher concentrations of hydrogen peroxide treatments may seem appealing for more severe cHABs but there is currently little understanding of the environmental impacts of this approach. Of specific concern is the associated microbial community, which may play key roles in the succession/recovery process post-treatment. To better understand impacts of a high concentration treatment on non-target microbial communities, we applied a hydrogen peroxide spray equating to a total volume concentration of 14 mM (473 mg/L, 0.04%) to 250 L mesocosms containing Microcystis bloom biomass, monitoring treatment and control mesocosms for 4 days. Cyanobacteria dominated control mesocosms throughout the experiment while treatment mesocosms experienced a 99% reduction, as determined by bacterial amplicon sequencing, and a 92% reduction in bacterial cell density within 1 day post-treatment. Only the bacterial community exhibited signs of regrowth, with a fold change of 9.2 bacterial cell density from day 1 to day 2. Recovery consisted of succession by Planctomycetota (47%) and Gammaproteobacteria (17%), which were likely resilient due to passive cell component compartmentalization and rapid upregulation of dnaK and groEL oxidative stress genes, respectively. The altered microbiome retained beneficial functionality of microcystin degradation through a currently recognized but unidentified pathway in Gammaproteobacteria, resulting in a 70% reduction coinciding with bacterial regrowth. There was also an 81% reduction of both total nitrogen and phosphorus, as compared to 91 and 93% in the control, respectively, due to high expressions of genes related to nitrogen (argH, carB, glts, glnA) and phosphorus (pntAB, phoB, pstSCB) cycling. Overall, we found a portion of the bacterial community was resilient to the high-concentration hydrogen peroxide treatment, resulting in Planctomycetota and Gammaproteobacteria dominance. This high-concentration treatment may be suitable to rapidly end cHABs which have already negatively impacted the aquatic environment rather than allow them to persist., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

The global increase in harmful algal blooms (HABs) has become a growing concern over the years, and New York State (NYS) is no exception. The Finger Lakes region in NYS has been identified as a hotspot for HABs, with Cayuga Lake having the highest number of blooms reported. The Cayuga Lake HABs Monitoring Program has been tracking cHABs (dominant bloom taxa, chlorophyll A, and microcystin levels) since 2018. However, limited research has been conducted on the microbiome of HABs in this region. In this study, the microbiome of HABs in the Cayuga Lake was surveyed and compared with non-HAB baseline samples. Using 16S rDNA community analysis, common bloom-forming cyanobacteria, were identified, with Microcystis being the dominant taxa in high toxin blooms. Further, this study evaluated the ability of Microcystis mcyA qPCR to detect elevated levels of potential toxigenic Microcystis in water samples using both benchtop and handheld qPCR devices. The results showed good performance of the qPCR assay as a screening for high toxin versus low/no toxin blooms. Additionally, the handheld qPCR device holds potential for in-field rapid (<1 h) screenings for high toxin blooms. This study provides insights into the microbiome of HABs in Cayuga Lake and offers a potential tool for rapid screening of high toxin blooms., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

Harmful algal blooms (HABs) are a threat to freshwater systems over the world due to the production of hepatotoxins like microcystin (MC), and nuisance taste and odour (T&O) compounds like 2-methylisoborneol (MIB). While MCs are known to cause detrimental effects to both water quality and human health, MIB is only reported to cause aesthetical problems. In this study, we investigated a tropical, urban lake that was experiencing persistent MC and MIB events. Although it was dominated by Microcystis blooms, analysis revealed that the toxigenic Microcystis were not the only species driving the MC concentrations. Additionally, there was also a lack of causative species for the MIB events. Through isolation, we have identified three toxigenic Microcystis found to produce four different variants of MCs, and two novel non-toxigenic Microcystis that were capable of producing MIB. The ability to produce MIB had never been previously reported for this species. Compared to other major producers such as Planktothricoides sp. and Streptomyces sp., the MIB synthase genes of our Microcystis sp. strains were partial, illustrating the possibility of unique synthesis pathways. The Microcystis sp. strains were found to produce about 2.77-5.22 fg MIB cell -1 , with a majority of the contents (70-80 %) existing in the extracellular phase. Correlation analysis of field study indicated that phosphorus limitation may have an indirect effect on non-toxigenic Microcystis abundance and proportion by influencing the toxigenic genotype, suggesting that current measures to control HABs may favour the proliferation of the non-toxigenic Microcystis. The potential for Microcystis sp. to produce MIB through unique synthesis pathway, coupled with the potential dominance of non-toxigenic genotypes in Microcystis blooms, signals the possibility that non-toxigenic Microcystis should be monitored as well., Competing Interests: Declaration of competing interest All authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Superior antagonistic activity against axenic Microcystis aeruginosa PCC7806 was observed with Paucibacter sp. B51 isolated from cyanobacterial bloom samples among 43 tested freshwater bacterial species. Complete genome sequencing, analyzing average nucleotide identity and digital DNA-DNA hybridization, designated the B51 strain as Paucibacter aquatile. Electron and fluorescence microscopic image analyses revealed the presence of the B51 strain in the vicinity of M. aeruginosa cells, which might provoke direct inhibition of the photosynthetic activity of the PCC7806 cells, leading to perturbation of cellular metabolisms and consequent cell death. Our speculation was supported by the findings that growth failure of the PCC7806 cells led to low pH conditions with fewer chlorophylls and down-regulation of photosystem genes (e.g., psbD and psaB) during their 48-h co-culture condition. Interestingly, the concentrated ethyl acetate extracts obtained from B51-grown supernatant exhibited a growth-inhibitory effect on PCC7806. The physical separation of both strains by a filter system led to no inhibitory activity of the B51 cells, suggesting that contact-mediated anti-cyanobacterial compounds might also be responsible for hampering the growth of the PCC7806 cells. Bioinformatic tools identified 12 gene clusters that possibly produce secondary metabolites, including a class II lasso peptide in the B51 genome. Further chemical analysis demonstrated anti-cyanobacterial activity from fractionated samples having a rubrivinodin-like lasso peptide, named paucinodin. Taken together, both contact-mediated inhibition of photosynthesis and the lasso peptide secretion of the B51 strain are responsible for the anti-cyanobacterial activity of P. aquatile B51., (© 2023 Phycological Society of America.)

Microcystis-dominated cyanobacterial harmful algal blooms (cyanoHABs) have a global impact on freshwater environments, affecting both wildlife and human health. Microcystis diversity and function in field samples and laboratory cultures can be determined by sequencing whole genomes of cultured isolates or natural populations, but these methods remain computationally and financially expensive. Amplicon sequencing of marker genes is a lower cost and higher throughput alternative to characterize strain composition and diversity in mixed samples. However, the selection of appropriate marker gene region(s) and primers requires prior understanding of the relationship between single gene genotype, whole genome content, and phenotype. To identify phylogenetic markers of Microcystis strain diversity, we compared phylogenetic trees built from each of 2,351 individual core genes to an established phylogeny and assessed the ability of these core genes to predict whole genome content and bioactive compound genotypes. We identified single-copy core genes better able to resolve Microcystis phylogenies than previously identified marker genes. We developed primers suitable for current Illumina-based amplicon sequencing with near-complete coverage of available Microcystis genomes and demonstrate that they outperform existing options for assessing Microcystis strain composition. Results showed that genetic markers can be used to infer Microcystis gene content and phenotypes such as potential production of bioactive compounds , although marker performance varies by bioactive compound gene and sequence similarity. Finally, we demonstrate that these markers can be used to characterize the Microcystis strain composition of laboratory or field samples like those collected for surveillance and modeling of Microcystis-dominated cyanobacterial harmful algal blooms., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Cyanobacterial blooms pose a global environmental concern, with various genera contributing to their formation. The harmfulness of cyanobacterial blooms varies depending on the specific genus, yet the factors triggering their formation remain incompletely understood. This study conducted qPCR of sediment DNA in Lake Erhai to reconstruct the historical succession of three common bloom-forming cyanobacterial genera (i.e., Microcystis, Dolichospermum, and Aphanizomenon). The driving factors and their corresponding thresholds were identified, and human activities related to driving factors were evaluated. The results revealed two successions in the past century. The first succession transitioned from Aphanizomenon (1902-1978) to Microcystis and Dolichospermum (1978-1999), driven by TN:TP and TP. The second succession shifted from Microcystis and Dolichospermum (1978-1999) to Microcystis (1999-2010), driven by TP, TN:TP, and temperature. The thresholds of TP and TN:TP for the Microcystis bloom were 0.023 mg/L and 17, respectively. TN:TP was significantly influenced by domestic pollution and crop farming in both successions, while TP was significantly impacted by domestic pollution in the first succession and by pollution from crop and dairy farming in the second succession. These results shed light on the underlying mechanism responsible for the blooms of various cyanobacterial genera and could serve as a valuable reference for effectively preventing and controlling nutrient input in the watershed., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Shao Yang reports financial support was provided by National Science and Technology Major Project , China. Shao Yang reports was provided by National Natural Science Foundation, China., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Toxigenic Microcystis blooms periodically disrupt the stabilization ponds of wastewater treatment plants (WWTPs). Dense proliferations of Microcystis cells within the surface waters (SWs) impede the water treatment process by reducing the treatment efficacy of the latent WWTP microbiome. Further, water quality is reduced when conventional treatment leads to Microcystis cell lysis and the release of intracellular microcystins into the water column. Recurrent seasonal Microcystis blooms cause significant financial burdens for the water industry and predicting their source is vital for bloom management strategies. We investigated the source of recurrent toxigenic Microcystis blooms at Australia's largest lagoon-based municipal WWTP in both sediment core (SC) and SW samples between 2018 and 2020. Bacterial community composition of the SC and SW samples according to 16S rRNA gene amplicon sequencing showed that Microcystis sp. was dominant within SW samples throughout the period and reached peak relative abundances (32%) during the summer. The same Microcystis Amplicon sequence variants were present within the SC and SW samples indicating a potential migratory population that transitions between the sediment water and SWs during bloom formation events. To investigate the potential of the sediment to act as a repository of viable Microcystis cells for recurrent bloom formation, a novel in-vitro bloom model was established featuring sediments and sterilized SW collected from the WWTP. Microcystin-producing Microcystis blooms were established through passive resuspension after 12 weeks of incubation. These results demonstrate the capacity of Microcystis to transition between the sediments and SWs in WWTPs, acting as a perennial inoculum for recurrent blooms.IMPORTANCECyanobacterial blooms are prevalent to wastewater treatment facilities owing to the stable, eutrophic conditions. Cyanobacterial proliferations can disrupt operational procedures through the blocking of filtration apparatus or altering the wastewater treatment plant (WWTP) microbiome, reducing treatment efficiency. Conventional wastewater treatment often results in the lysis of cyanobacterial cells and the release of intracellular toxins which pose a health risk to end users. This research identifies a potential seeding source of recurrent toxigenic cyanobacterial blooms within wastewater treatment facilities. Our results demonstrate the capacity of Microcystis to transition between the sediments and surface waters (SWs) of wastewater treatment ponds enabling water utilities to develop adequate monitoring and management strategies. Further, we developed a novel model to demonstrate benthic recruitment of toxigenic Microcystis under laboratory conditions facilitating future research into the genetic mechanisms behind bloom development., Competing Interests: The authors declare no conflict of interest.

Microcystis blooms have a marked effect on microbial taxonomical diversity in eutrophic lakes, but their influence on the composition of microbial functional genes is still unclear. In this study, the free-living microbial functional genes (FMFG) composition was investigated in the period before Microcystis blooms (March) and during Microcystis blooms (July) using a comprehensive functional gene array (GeoChip 5.0). The composition and richness of FMFG in the water column was significantly different between these two periods. The FMFG in March was enriched in the functional categories of nitrogen, sulfur, and phosphorus cycling, whereas the FMFG in July was enriched in carbon cycling, organic remediation, and metal homeostasis. Molecular ecological network analysis further demonstrated fewer functional gene interactions and reduced complexity in July than in March. Module hubs of the March network were mediated by functional genes associated with carbon, nitrogen, sulfur, and phosphorus, whereas those in July by a metal homeostasis functional gene. We also observed stronger deterministic processes in the FMFG assembly in July than in March. Collectively, this study demonstrated that Microcystis blooms induced significant changes in FMFG composition and metabolic potential, and abundance-information, which can support the understanding and management of biogeochemical cycling in eutrophic lake ecosystems., (© The Author(s) 2023. Published by Oxford University Press on behalf of FEMS.)

Since the 2007 water crisis occurred in Lake Taihu, substantial measures have been taken to restore the lake. This study evaluates the effectiveness of these restoration activities. We examined the physicochemical parameters and the distribution of microcystin and Microcystis in both the water column and sediment during the bloom period of May 2020 to October 2020. The mean value of extracellular and intracellular microcystin content was 0.12 μg L -1 and 16.26 μg L -1 , respectively. The mean value of microcystin in sediment was 172.02 ng g -1 and peaked in August. The concentration in the water and sediment was significantly lower than the historical average concentration. The abundance of toxigenic Microcystis and total Microcystis in the water column ranged from 2.61 × 10 2 to 2.25 × 10 9 copies·L -1 and 8.28 × 10 5 to 2.76 × 10 9 copies·L -1 , respectively. The proportion of toxic Microcystis in the sediment ranging from 31.2% to 19.12%. The highest and lowest region was Meiliang Bay and Grass-algae type zone, respectively. The copy number of the 16S rRNA gene was 1-4 orders of magnitude higher than that of mcyA gene in populations of Microcystis, indicating that non-toxic Microcystis was the dominant form in the majority of the lake. The abundance of toxic Microcystis in the water column was positively correlated with total phosphorus, PO 4 3- -P and pH, while the water temperature played distinct role to the distribution of toxic Microcystis in sediment. Our research indicated phosphorus remains a key factor influencing the toxic Microcystis and microcystins in the water column. pH played distinct roles in the distribution of microcystins in sediment and water column. The increasing water temperature is a threat. Explicit management actions and policies, which take into account nutrient concentrations, pH, and increasing temperatures, are necessary to understand and control the distribution of microcystin and Microcystis in Lake Taihu., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

A novel alphaproteobacterial strain JXJ CY 41 T was isolated from a culture mass of Microcystis, collected from Lake Dianchi, south-west, China. Strain JXJ CY 41 T was gram-strain-negative, aerobic, motile, with rod-shaped cells (0.4-1.0 × 1.7-3.5 μm). It was positive for catalase and starch hydrolysis, negative for oxidase and hydrolysis of Tweens (20, 40, and 80). Growth occurred at 10-44 °C, pH 5.0-10.0, and 0-5.0% (w/v) NaCl. Major fatty acids included C 16:0 (28.1%), 11-methyl C 18:1 ω7c (36.7%) and C 18:1 ω7c (20.8%). Q10 was the sole ubiquinone. The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, glycolipid, and an unidentified lipid. The DNA G + C content was 63.1%. Its 16S rRNA gene sequence showed high similarities with Devosia oryziradicis G19 T (99.5%; not validly published), D. yakushimensis Yak96B T (98.3%) and D. ginsengisoli Gsoil 520 T (98.1%), and less than 98.1% similarities with other members of the genus Devosia. The digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values between strain JXJ CY 41 T and its 5 closest similar strains were 19.9-24.1% and 75.7-80.5%, respectively. Based on the data above, strain JXJ CY 41 T was identified as a novel species of the genus Devosia, for which the epithet Devosia lacusdianchii sp. nov. was proposed. The type strain is JXJ CY 41 T (= KCTC 72812 T  = CGMCC 1.17502 T ). Strain JXJ CY 41 T exhibited different interactions with Microcystis aeruginosa FACHB-905 (Maf) under different conditions, and Maf could control the bacterial cellular density by secreting unknown specific chemical compounds according to its nutritional requirements., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Early monitoring of Microcystis, a cyanobacterium that produces microcystin, is paramount in order to confirm the presence of Microcystis spp. Both phenotypic and genotypic methods have been used. The phenotypic methods provide the presence of the microcystis but do not confirm its species type and toxin produced. Additionally, phenotypic methods cannot differentiate toxigenic from non-toxigenic Microcystis. Therefore, the current protocol also describes genetic methods based on PCR to detect toxigenic Microcystis spp. based on microcystin synthetase E (mcy E) gene and 16-23S RNA genes for species-specific identification, which can effectively comprehend distinct lineages and discrimination of potential complexity of microcystin populations. The presence of these microcystin toxins in blood, in most cases, indicates contamination of drinking water by cyanobacteria. The methods presented herein are used to identify microcystin toxins in drinking water and blood., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Microcystis aeruginosa (M. aeruginosa) causes massive blooms in eutrophic freshwater and releases microcystin. Poyang Lake is the largest freshwater lake in China and has kept a mid-nutrient level in recent years. However, there is little research on microcystin production in Poyang Lake. In this study, water and sediment samples from ten sampling sites in Poyang Lake were collected from May to December in 2020, and from January to April in 2021 respectively. Microcystis genes (mcyA, mcyB, 16 s rDNA) were quantified by real-time fluorescence quantitative PCR analysis, and then the spatial and temporal variation of mcy genes, physicochemical factors, and bacterial population structure in the lake was analyzed. The relationship between the abundance of mcy genes and physicochemical factors in water column was also revealed. Results indicated that the microcystin-producing genes mcyA and mcyB showed significant differences in spatial and temporal levels as well, which is closely related to the physicochemical factors especially the water temperature (p < 0.05) and the nitrogen content (p < 0.05). The abundance of mcy genes in the sediment in December affected the abundance of mcy genes in the water column in the next year, while the toxic Microcystis would accumulate in the sediment. In addition to the toxic Microcystis, we also found a large number of non-toxic Microcystis in the water column and sediment, and the ratio of toxic to non-toxic species can also affect the toxicity production of M. aeruginosa. Overall, the results showed that M. aeruginosa toxin-producing genes in Poyang Lake distributed spatially and temporally which related to the physicochemical factors of Poyang Lake., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Many freshwater cyanobacteria, including Microcystis aeruginosa, lack several known antibiotic resistance genes; however, both axenic and xenic M. aeruginosa strains exhibited high antibiotic resistance against many antibiotics under our tested concentrations, including colistin, trimethoprim, and kanamycin. Interestingly, axenic PCC7806, although not the xenic NIBR18 and NIBR452 strains, displayed susceptibility to ampicillin and amoxicillin, indicating that the associated bacteria in the phycosphere could confer such antibiotic resistance to xenic strains. Fluorescence and scanning electron microscopic observations revealed their tight association, leading to possible community-level β-lactamase activity. Combinatory treatment of ampicillin with a β-lactamase inhibitor, sulbactam, abolished the ampicillin resistance in the xenic stains. The nitrocefin-based assay confirmed the presence of significant community-level β-lactamase activity. Our tested low ampicillin concentration and high β-lactamase activity could potentially balance the competitive advantage of these dominant species and provide opportunities for the less competitive species, thereby resulting in higher bacterial diversity under ampicillin treatment conditions. Non-PCR-based metagenome data from xenic NIBR18 cultures revealed the dominance of bla OXA -related antibiotic resistance genes followed by other class A β-lactamase genes (AST-1 and FAR-1). Alleviation of ampicillin toxicity could be observed only in axenic PCC7806, which had been cocultured with β-lactamase from other freshwater bacteria. Our study suggested M. aeruginosa develops resistance to old-class β-lactam antibiotics through altruism, where associated bacteria protect axenic M. aeruginosa cells., (© 2023. The Author(s), under exclusive licence to Microbiological Society of Korea.)

Background: Numerous studies in the past have expanded our understanding of the genetic differences of global distributed cyanobacteria that originated around billions of years ago, however, unraveling how gene gain and loss drive the genetic evolution of cyanobacterial species, and the trade-off of these evolutionary forces are still the central but poorly understood issues., Results: To delineate the contribution of gene flow in mediating the hereditary differentiation and shaping the microbial evolution, a global genome-wide study of bloom-forming cyanobacterium, Microcystis aeruginosa species complex, provided robust evidence for genetic diversity, reflected by enormous variation in gene repertoire among various strains. Mathematical extrapolation showed an 'open' microbial pan-genome of M. aeruginosa species, since novel genes were predicted to be introduced after new genomes were sequenced. Identification of numerous horizontal gene transfer's signatures in genome regions of interest suggested that genome expansion via transformation and phage-mediated transduction across bacterial lineage as an evolutionary route may contribute to the differentiation of Microcystis functions (e.g., carbohydrate metabolism, amino acid metabolism, and energy metabolism). Meanwhile, the selective loss of some dispensable genes at the cost of metabolic versatility is as a mean of adaptive evolution that has the potential to increase the biological fitness., Conclusions: Now that the recruitment of novel genes was accompanied by a parallel loss of some other ones, a trade-off in gene content may drive the divergent differentiation of M. aeruginosa genomes. Our study provides a genetic framework for the evolution of M. aeruginosa species and illustrates their possible evolutionary patterns., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Denitrification is the most important process for nitrogen removal in eutrophic lakes and was mostly investigated in lake sediment. Denitrification could also be mediated by cyanobacterial aggregates, yet how this process impacts nitrogen (N) availability and the associated autotroph-heterotroph relationships within cyanobacterial aggregates has not been investigated. In this study, incubation experiments with nitrate amendment were conducted with Microcystis aggregates (MAs). Measurement of nitrogen contents, 16S rRNA-based microbial community profiling and metatranscriptomic sequencing were used to jointly assess nitrogen turnover dynamics, as well as changes in microbial composition and gene expression. Strong denitrification potential was revealed, and maximal N removal was achieved within two days, after which the communities entered a state of severe N limitation. Changes of active microbial communities were further promoted both with regard to taxonomic composition and transcriptive activities. Expression of transportation-related genes confirmed competition for N sources by Microcystis and phycospheric communities. Strong stress response to reactive oxygen species by Microcystis was revealed. Notably, interspecific relationships among Microcystis and phycospheric communities exhibited a shift toward antagonistic interactions, particularly evidenced by overall increased expression of genes related to cell lysis and utilization of cellular materials. Patterns of fatty acid and starch metabolism also suggested changes in carbon metabolism and cross-feeding patterns within MAs. Taken together, this study demonstrated substantial denitrification potential of MAs, which, importantly, further induced changes in both metabolic activities and autotroph-heterotroph interactions. These findings also highlight the key role of nutrient condition in shaping autotroph-heterotroph relationships., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Cyanobacteria can dominate the algal community in wastewater ponds, which can lead to the production of cyanotoxins and their release into the environment. We applied traditional and molecular techniques to identify cyanotoxin hazards and high-risk periods in a tropical wastewater treatment system. Potentially toxic cyanobacteria were identified by microscopy and amplicon sequencing over the course of a year. Toxin gene levels were monitored and compared to toxin production to identify likely toxin producing species and high-risk periods. Cyanobacteria were persistent in the effluent year-round, with Planktothrix and Microcystis the most abundant genera; Microcystis could not be resolved beyond genus using amplicon sequencing, but M. flos-aquae was identified as a dominant species by microscopy. Microcystin toxin was detected for the first time in treated effluent at the beginning of the wet season (December 2018), which correlated with an increase in Microcystis amplicon sequence abundance and elevated microcystin toxin gene (mcyE/ndaF) levels. Concomitantly, microscopy data showed an increase in M. flos-aquae but not M. aeruginosa. These data informed a refined sampling campaign in 2019 and results showed a strong correlation between mcyE/ndaF gene abundance, microcystin toxin levels and Microcystis amplicon sequence abundance. Microscopy data showed that in addition to M. flos-aquae, M. aeruginosa was also abundant in February and March 2019, with highest levels coinciding with toxin detection and toxin gene levels. M. aeruginosa was the most abundant Microcystis species detected in selected treated effluent samples by metagenomics analysis, and elevated levels coincided with toxin production. All microcystin genes in the biosynthesis pathway were detected, but microcystin genes from Planktothrix agardhii were not detected. Gene toxin assays were successfully used to predict microcystin production in this wastewater system. Changes in amplicon sequence relative abundance were a useful indicator of changes in the cyanobacterial community. We found that metagenomics was useful not just for identifying the most abundant Microcystis species, but the detection of microcystin biosynthesis genes helped confirm this genus as the most likely toxin producer in this system. We recommend toxin gene testing for the early detection of potential toxin producing cyanobacteria to manage the risk of toxicity and allow the implementation of risk management strategies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

α-Ketoglutarate decarboxylase is a crucial enzyme in the tricarboxylic acid cycle of cyanobacteria, catalyzing the non-oxidative decarboxylation of α-ketoglutarate to produce succinate semialdehyde and CO 2 . The decarboxylation process is reliant on the cofactor of thiamine diphosphate. However, this enzyme's biochemical and structural properties have not been well characterized. In this work, two α-ketoglutarate decarboxylases encoded by MAE&#95;06010 and MiAbw&#95;01735 genes from Microcystis aeruginosa NIES-843 (MaKGD) and NIES-4325 (MiKGD), respectively, were overexpressed and purified by using an Escherichia coli expression system. It was found that MaKGD exhibited 9.2-fold higher catalytic efficiency than MiKGD, which may be attributed to the absence of glutamate decarboxylase in Microcystis aeruginosa NIES-843. Further biochemical investigation of MaKGD demonstrated that it displayed optimum activity at pH 6.5-7.0 and was most activated by Mg 2+ . Additionally, MaKGD showed substrate specificity towards α-ketoglutarate. Structural modeling and autodocking results revealed that the active site of MaKGD contained a distinct binding pocket where α-ketoglutarate and thiamine diphosphate interacted with specific amino acid residues via hydrophobic interactions, hydrogen bonds and salt bridges. Furthermore, the mutagenesis study provided strong evidence supporting the importance of certain residues in the catalysis of MaKGD. These findings provide new insights into the structure-function relationships of α-ketoglutarate decarboxylases from cyanobacteria.

This study aimed to test the hypothesis of the year-round presence of toxigenic Microcystis and cyanotoxins in the water and ice of the shallow eutrophic Lake Ytyk-Kyuyol located in the continuous permafrost zone. Three independent approaches-mass-spectrometry, molecular methods and light microscopy-were applied in the study. The cyanobacterial biomass ranged from 1.0 × 10 -4 to 4.8 mg L -1 . Microcystis flos-aquae and M. aeruginosa were the dominant morphospecies in plankton throughout the observation. In environmental DNA, the presence of M. aeruginosa was supported and mcy gene regions responsible for microcystin biosynthesis were detected through a BLAST (Basic Local Alignment Search Tool) search and phylogenetic estimation based on newly obtained 16S rRNA, 16S-23S ITS rRNA, mcy A and mcy E nucleotide sequences. The intracellular microcystin concentration ranged from <0.1 to 803 ng L -1 , and the microcystin quota in the Microcystis biomass was extremely low. For the first time, it was shown that Microcystis cells containing mcy genes and microcystins presented permanently in the water column, both during the ice-free period and under ice, as well as inside thick ice covers within 7 months of severe winter. We hypothesized that minor pelagic and ice populations of Microcystis could participate in increasing cell density in the spring. However, further studies are needed to confirm the viability of the overwintering Microcystis colonies in the water and inside the ice of Lake Ytyk-Kyuyol.

Cyanobacterial harmful algal blooms (cyanoHABs) dominated by Microcystis spp. have significant public health and economic implications in freshwater bodies around the world. These blooms are capable of producing a variety of cyanotoxins, including microcystins, that affect fishing and tourism industries, human and environmental health, and access to drinking water. In this study, we isolated and sequenced the genomes of 21 primarily unialgal Microcystis cultures collected from western Lake Erie between 2017 and 2019. While some cultures isolated in different years have a high degree of genetic similarity (genomic Average Nucleotide Identity >99%), genomic data show that these cultures also represent much of the breadth of known Microcystis diversity in natural populations. Only five isolates contained all the genes required for microcystin biosynthesis while two isolates contained a previously described partial mcy operon. Microcystin production within cultures was also assessed using Enzyme-Linked Immunosorbent Assay (ELISA) and supported genomic results with high concentrations (up to 900 μg L⁻¹) in cultures with complete mcy operons and no or low toxin detected otherwise. These xenic cultures also contained a substantial diversity of bacteria associated with Microcystis, which has become increasingly recognized as an essential component of cyanoHAB community dynamics. These results highlight the genomic diversity among Microcystis strains and associated bacteria in Lake Erie, and their potential impacts on bloom development, toxin production, and toxin degradation. This culture collection significantly increases the availability of environmentally relevant Microcystis strains from temperate North America., Competing Interests: Declaration of Competing Interest The authors declare no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Bloom-forming phytoplankton dynamics are still unpredictable, even though it is known that several abiotic factors, such as nutrient availability and temperature, are key factors for bloom development. We investigated whether biotic factors, i.e. the bacterioplankton composition (via 16SrDNA metabarcoding), were correlated with phytoplankton dynamics, through a weekly monitoring of a shallow lake known to host recurrent cyanobacterial blooms. We detected concomitant changes in both bacterial and phytoplankton community biomass and diversity. During the bloom event, a significant decrease in phytoplankton diversity, was detected, with a first co-dominance of Ceratium, Microcystis and Aphanizomenon, followed by a co-dominance of the two cyanobacterial genera. In the same time, we observed a decrease of the particle-associated (PA) bacterial richness and the emergence of a specific bacterial consortium that was potentially better adapted to the new nutritional niche. Unexpectedly, changes in PA bacterial communities occurred just before the development the emergence of the phytoplanktonic bloom and the associated modification of the phytoplanktonic community composition, suggesting that changes in environmental conditions leading to the bloom, were first sensed by the bacterial PA community. This last was quite stable throughout the bloom event, even though there were changes in the blooming species, suggesting that the association between cyanobacterial species and bacterial communities may not be as tight as previously described for monospecific blooming communities. Finally, the dynamics of the free-living (FL) bacterial communities displayed a different trajectory from those of the PA and phytoplankton communities. This FL communities can be viewed as a reservoir for bacterial recruitment for the PA fraction. Altogether, these data also highlight s that the spatial organization within these different microenvironments in the water column is a relevant factor in the structuring of these communities., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Background: Phosphonates are the main components in the global phosphorus redox cycle. Little is known about phosphonate metabolism in freshwater ecosystems, although rapid consumption of phosphonates has been observed frequently. Cyanobacteria are often the dominant primary producers in freshwaters; yet, only a few strains of cyanobacteria encode phosphonate-degrading (C-P lyase) gene clusters. The phycosphere is defined as the microenvironment in which extensive phytoplankton and heterotrophic bacteria interactions occur. It has been demonstrated that phytoplankton may recruit phycospheric bacteria based on their own needs. Therefore, the establishment of a phycospheric community rich in phosphonate-degrading-bacteria likely facilitates cyanobacterial proliferation, especially in waters with scarce phosphorus. We characterized the distribution of heterotrophic phosphonate-degrading bacteria in field Microcystis bloom samples and in laboratory cyanobacteria "phycospheres" by qPCR and metagenomic analyses. The role of phosphonate-degrading phycospheric bacteria in cyanobacterial proliferation was determined through coculturing of heterotrophic bacteria with an axenic Microcystis aeruginosa strain and by metatranscriptomic analysis using field Microcystis aggregate samples., Results: Abundant bacteria that carry C-P lyase clusters were identified in plankton samples from freshwater Lakes Dianchi and Taihu during Microcystis bloom periods. Metagenomic analysis of 162 non-axenic laboratory strains of cyanobacteria (consortia cultures containing heterotrophic bacteria) showed that 20% (128/647) of high-quality bins from eighty of these consortia encode intact C-P lyase clusters, with an abundance ranging up to nearly 13%. Phycospheric bacterial phosphonate catabolism genes were expressed continually across bloom seasons, as demonstrated through metatranscriptomic analysis using sixteen field Microcystis aggregate samples. Coculturing experiments revealed that although Microcystis cultures did not catabolize methylphosphonate when axenic, they demonstrated sustained growth when cocultured with phosphonate-utilizing phycospheric bacteria in medium containing methylphosphonate as the sole source of phosphorus., Conclusions: The recruitment of heterotrophic phosphonate-degrading phycospheric bacteria by cyanobacteria is a hedge against phosphorus scarcity by facilitating phosphonate availability. Cyanobacterial consortia are likely primary contributors to aquatic phosphonate mineralization, thereby facilitating sustained cyanobacterial growth, and even bloom maintenance, in phosphate-deficient waters. Video Abstract., (© 2023. The Author(s).)

The majority of bacterial antagonists identified to date are active against Microcystis. Therefore, this study aimed to isolate and characterize novel cyanolytic bacterial strains antagonistic against bloom-forming filamentous cyanobacteria. The bacterial strain BG-E isolated from the Bandagiriya Wewa in Sri Lanka was identified as Pseudomonas fluorescens (MZ007859) based on the 16S rRNA gene sequencing. BG-E showed 82% and 73% cyanolytic activity (CA) against Pseudanabaena sp. LW2 (MW288948) and Pseudanabaena lonchoides LW1 (MW288940), respectively, after 10 days of inoculation. The light microscopic images affirmed the complete disintegration in the filamentous structures of the tested Pseudanabaena species. The bacterial cell density of 15% v/v showed the CA with 95% and 89% cell lysis, respectively, in P. lonchoides and Pseudanabaena sp. LW2. Moreover, the results showed that >50% CA could be achieved by 0.100 and 1.00 (OD 730 ) cell densities for these same species. The highest CA of the cell-free supernatant of BG-E against P. lonchoides and bacterial culture against Pseudanabaena sp. LW2 illustrated the species-specific mode of action of BG-E. Although BG-E efficiently lysed the tested cyanobacterial species, the results of the MC-biodegradation assay confirmed its inability to degrade MC-LR cyanotoxin. Further, the BG-E strain lacks the mlrABCD gene cluster which is known to be responsible for the enzymatic degradation of MCs. The overall findings highlighted the applicability of P. fluorescens BG-E as a biological controlling agent to terminate blooms of freshwater filamentous cyanobacteria genus Pseudanabaena. The incorporation of cyanotoxin-degrading heterotrophic bacteria is recommended as a means of controlling toxic Pseudanabaena blooms., (© 2023 Phycological Society of America.)

Microcystis spp. produce diverse secondary metabolites within freshwater cyanobacterial harmful algal blooms (cyanoHABs) around the world. In addition to the biosynthetic gene clusters (BGCs) encoding known compounds, Microcystis genomes harbor numerous BGCs of unknown function, indicating a poorly understood chemical repertoire. While recent studies show that Microcystis produces several metabolites in the lab and field, little work has focused on analyzing the abundance and expression of its broader suite of BGCs during cyanoHAB events. Here, we use metagenomic and metatranscriptomic approaches to track the relative abundance of Microcystis BGCs and their transcripts throughout the 2014 western Lake Erie cyanoHAB. The results indicate the presence of several transcriptionally active BGCs that are predicted to synthesize both known and novel secondary metabolites. The abundance and expression of these BGCs shifted throughout the bloom, with transcript abundance levels correlating with temperature, nitrate, and phosphorus concentrations and the abundance of co-occurring predatory and competitive eukaryotic microorganisms, suggesting the importance of both abiotic and biotic controls in regulating expression. This work highlights the need for understanding the chemical ecology and potential risks to human and environmental health posed by secondary metabolites that are produced but often unmonitored. It also indicates the prospects for identifying pharmaceutical-like molecules from cyanoHAB-derived BGCs. IMPORTANCE Microcystis spp. dominate cyanobacterial harmful algal blooms (cyanoHABs) worldwide and pose significant threats to water quality through the production of secondary metabolites, many of which are toxic. While the toxicity and biochemistry of microcystins and several other compounds have been studied, the broader suite of secondary metabolites produced by Microcystis remains poorly understood, leaving gaps in our understanding of their impacts on human and ecosystem health. We used community DNA and RNA sequences to track the diversity of genes encoding synthesis of secondary metabolites in natural Microcystis populations and assess patterns of transcription in western Lake Erie cyanoHABs. Our results reveal the presence of both known gene clusters that encode toxic secondary metabolites as well as novel ones that may encode cryptic compounds. This research highlights the need for targeted studies of the secondary metabolite diversity in western Lake Erie, a vital freshwater source to the United States and Canada., Competing Interests: The authors declare no conflict of interest.