A novel hierarchical network-based approach to unveil the complexity of functional microbial genome.

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.

Mobilome impacts on physiology in the widely used non-toxic mutant Microcystis aeruginosa PCC 7806 ΔmcyB and toxic wildtype.

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.

Endogenous Production and Vibrational Analysis of Heavy-Isotope-Labeled Peptides from Cyanobacteria.

Stable isotope labeling is an extremely useful tool for characterizing the structure, tracing the metabolism, and imaging the distribution of natural products in living organisms using mass-sensitive measurement techniques. In this study, a cyanobacterium was cultured in 15 N/13 C-enriched media to endogenously produce labeled, bioactive oligopeptides. The extent of heavy isotope incorporation in these peptides was determined with LC-MS, while the overall extent of heavy isotope incorporation in whole cells was studied with nanoSIMS and AFM-IR. Up to 98 % heavy isotope incorporation was observed in labeled cells. Three of the most abundant peptides, microcystin-LR (MCLR), cyanopeptolin-A (CYPA), and aerucyclamide-A (ACAA), were isolated and further studied with Raman and FTIR spectroscopies and DFT calculations. This revealed several IR and Raman active vibrations associated with functional groups not common in ribosomal peptides, like diene, ester, thiazole, thiazoline, and oxazoline groups, which could be suitable for future vibrational imaging studies. More broadly, this study outlines a simple and relatively inexpensive method for producing heavy-labeled natural products. Manipulating the bacterial culture conditions by the addition of specific types and amounts of heavy-labeled nutrients provides an efficient means of producing heavy-labeled natural products for mass-sensitive imaging studies.

Microbiome analysis reveals Microcystis blooms endogenously seeded from benthos within wastewater maturation ponds.

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.

Pseudomonas ZY-1 and Bacillus FY-1 protecting the rice seedlings from the harm of Pseudomonas aeruginosa via indirect seawead lysis.

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 106 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.

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

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

Application of microbubble air flotation to harvest Microcystis sp. from agriculture wastewater: The regulation and mechanisms.

The harvesting of microalgae is the main bottleneck of its large-scale biomass production, and seeking an efficient, green, and low-cost microalgae harvesting technology is one of the urgent problems to be solved. Microbubble air flotation has been proven to be an effective measure, but the mechanisms of microbubbles-algal cell attachment are still unclear. In this study, microbubble air flotation was used as a harvesting method for Microcystis cultured in agricultural wastewater. The process mechanism of microbubble air flotation harvesting microalgae in wastewater was fully revealed from three aspects (the design of bubble formation, the adhesion law, and the recovery rate of microalgae under different working conditions). The results show that the length of the release pipe is the main factor affecting the proportion of microbubbles with a particle size of less than 50 µm. In the process of adhesion, when the particle size of microbubbles is 0.6-1.7 times the size of Microcystis, the adhesion efficiency of microbubbles to Microcystis is the highest. Under the conditions of pressure 0.45 MPa, gas-liquid ratio 5%, and release pipe length 100 cm, the harvesting performance of Microcystis was the best. Microbubble air flotation has better harvesting performance (63.5%, collection rate) of Microcystis with higher density. By understanding the mechanism of microbubble flotation, the technical parameters of microbubble flotation for harvesting energy microalgae are optimized to provide support for the development of efficient and low-cost devices and equipment for collecting microalgae.

Grazing preferences of three species of amoebae on cyanobacteria and green algae.

Twenty species/isolates of cyanobacteria and green algae were isolated from cyanobacterial bloom samples in lakes associated with the upper Qu'Appelle River drainage system in southern Saskatchewan, Canada. Three amoebae species (Cochliopodium sp., Vannella sp. and Vermamoeba vermiformis) were also isolated from one of these samples, and were subjected to grazing assays to determine which species of cyanobacteria or algae could potentially serve as a food source. Amoeba grazing rates were quantified based on the diameter of the plaque after 12 days on agar plate assays, and by estimation of the amoeba population growth rate from the rate of increase of plaque area. The common cyanobacterial bloom-formers Dolichospermum sp. and Aphanizomenon flos-aquae supported high growth rates for all three amoebae, while green algae, with the exception of one green alga/amoeba combination, did not support growth of the tested amoebae. Many of the cyanobacterial and algal isolates that did not support amoebae growth were ingested, suggesting that ingestion did not determine grazing success. Overall, while the cyanobacteria Dolichospermum sp. and Aphanizomenon flos-aquae were suitable food sources for the amoebae, the other cyanobacteria were grazed in an unpredictable manner, with some species/strains grazed by some amoebae and some species not grazed at all.

Repeated Exposure Enhanced Toxicity of Clarithromycin on Microcystis aeruginosa Versus Single Exposure through Photosynthesis, Oxidative Stress, and Energy Metabolism Shift.

Antibiotics are being increasingly detected in aquatic environments, and their potential ecological risk is of great concern. However, most antibiotic toxicity studies involve single-exposure experiments. Herein, we studied the effects and mechanisms of repeated versus single clarithromycin (CLA) exposure on Microcystis aeruginosa. The 96 h effective concentration of CLA was 13.37 µg/L upon single exposure but it reduced to 6.90 µg/L upon repeated exposure. Single-exposure CLA inhibited algal photosynthesis by disrupting energy absorption, dissipation and trapping, reaction center activation, and electron transport, thereby inducing oxidative stress and ultrastructural damage. In addition, CLA upregulated glycolysis, pyruvate metabolism, and the tricarboxylic acid cycle. Repeated exposure caused stronger inhibition of algal growth via altering photosynthetic pigments, reaction center subunits biosynthesis, and electron transport, thereby inducing more substantial oxidative damage. Furthermore, repeated exposure reduced carbohydrate utilization by blocking the pentose phosphate pathway, consequently altering the characteristics of extracellular polymeric substances and eventually impairing the defense mechanisms of M. aeruginosa. Risk quotients calculated from repeated exposure were higher than 1, indicating significant ecological risks. This study elucidated the strong influence of repeated antibiotic exposure on algae, providing new insight into antibiotic risk assessment.

Reactivity of Cyanobacteria Metabolites with Ozone: Multicompound Competition Kinetics.

Cyanobacterial blooms occur at increasing frequency and intensity, notably in freshwater. This leads to the introduction of complex mixtures of their products, i.e., cyano-metabolites, to drinking water treatment plants. To assess the fate of cyano-metabolite mixtures during ozonation, a novel multicompound ozone (O3) competition kinetics method was developed. Sixteen competitors with known second-order rate constants for their reaction with O3 ranging between 1 and 108 M-1 s-1 were applied to cover a wide range of the O3 reactivity. The apparent second-order rate constants (kapp,O3) at pH 7 were simultaneously determined for 31 cyano-metabolites. kapp,O3 for olefin- and phenol-containing cyano-metabolites were consistent with their expected reactivity (0.4-1.7 × 106 M-1 s-1) while kapp,O3 for tryptophan- and thioether-containing cyano-metabolites were significantly higher than expected (3.4-7.3 × 107 M-1 s-1). Cyano-metabolites containing these moieties are predicted to be well abated during ozonation. For cyano-metabolites containing heterocycles, kapp,O3 varied from <102 to 5.0 × 103 M-1 s-1, giving first insights into the O3 reactivity of this class of compounds. Due to lower O3 reactivities, heterocycle- and aliphatic amine-containing cyano-metabolites may be only partially degraded by a direct O3 reaction near circumneutral pH. Hydroxyl radicals, which are formed during ozonation, may be more important for their abatement. This novel multicompound kinetic method allows a high-throughput screening of ozonation kinetics.

Inhibition of Fusarium graminearum growth and spore germination by a Streptomyces amritsarensis strain capable of killing and growing on Microcystis scum.

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.

Antiviral discovery in toxic cyanobacteria: Low hanging fruit in the age of pandemics.

The power of novel vaccination technologies and their rapid development were elucidated clearly during the COVID-19 pandemic. At the same time, it also became clear that there is an urgent need to discover and manufacture new antivirals that target emerging viral threats. Toxic species of cyanobacteria produce a range of bioactive compounds that makes them good candidates for drug discovery. Nevertheless, few studies demonstrate the antiviral potential of cyanobacteria. This is partly due to the lack of specific and simple protocols designed for the rapid detection of antiviral activity in cyanobacteria and partly because specialized facilities for work with pathogenic viruses are few and far between. We therefore developed an easy method for the screening of cyanobacterial cultures for antiviral activity and used our private culture collection of non-pathogenic virus isolates to show that antiviral activity is a prominent feature in the cyanobacterium Microcystis aeruginosa. In this proof-of-concept study, we show that M. aeruginosa extracts from three different cyanobacterial strains delay infection of diatom-infecting single-stranded DNA and single-stranded RNA viruses by up to 2 days. Our work shows the ease with which cyanobacteria from culture collections can be screened for antiviral activity and highlights the potential of cyanobacteria as an excellent source for the discovery of novel antiviral compounds, warranting further investigation.

Changes in microcystin concentration in Lake Taihu, 13 years (2007-2020) after the 2007 drinking water crisis.

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 × 102 to 2.25 × 109 copies·L-1 and 8.28 × 105 to 2.76 × 109 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, PO43--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.

Impact of butylparaben on growth dynamics and microcystin-LR production in Microcystis aeruginosa.

The presence of butylparaben (BP), a prevalent pharmaceutical and personal care product, in surface waters has raised concerns regarding its impact on aquatic ecosystems. Despite its frequent detection, the toxicity of BP to the cyanobacterium Microcystis aeruginosa remains poorly understood. This study investigates the influence of BP on the growth and physiological responses of M. aeruginosa. Results indicate that low concentrations of BP (below 2.5 mg/L) have negligible effects on M. aeruginosa growth, whereas higher concentrations (5 mg/L and 10 mg/L) lead to significant growth inhibition. This inhibition is attributed to the severe disruption of photosynthesis, evidenced by decreased Fv/Fm values and chlorophyll a content. BP exposure also triggers the production of reactive oxygen species (ROS), resulting in elevated activity of antioxidant enzymes. Excessive ROS generation stimulates the production of microcystin-LR (MC-LR). Furthermore, lipid peroxidation and cell membrane damage indicate that high BP concentrations cause cell membrane rupture, facilitating the release of MC-LR into the environment. Transcriptome analysis reveals that BP disrupts energy metabolic processes, particularly affecting genes associated with photosynthesis, carbon fixation, electron transport, glycolysis, and the tricarboxylic acid cycle. These findings underscore the profound physiological impact of BP on M. aeruginosa and highlight its role in stimulating the production and release of MC-LR, thereby amplifying environmental risks in aquatic systems.

Concentration-dependent effects of spinetoram on nontarget freshwater microalgae: A comparative study on Chlorella vulgaris and Microcystis aeruginosa.

The rising global demand for agricultural products is leading to the widespread application of pesticides, such as spinetoram, resulting in environmental pollution and ecotoxicity to nontarget organisms in aquatic ecosystems. This research focused on assessing the toxicity of spinetoram at various concentrations (0, 0.01, 0.1, 0.5, 1.0, and 3.0 mg L-1) on two common freshwater microalgae, Chlorella vulgaris and Microcystis aeruginosa, to shed light on the ecotoxicological effects of insecticides. Our findings demonstrate that M. aeruginosa is more sensitive to spinetoram than is C. vulgaris, with a concentration-dependent reduction in the growth rate observed for M. aeruginosa, whereas only the highest concentration of spinetoram adversely affected C. vulgaris. At a concentration of 0.01 mg L-1, the growth rate of M. aeruginosa unexpectedly increased beginning on day 7, indicating a potential hormetic effect. Although initial exposure to spinetoram improved the photosynthetic efficiency of both microalgae strains at all concentrations, detrimental effects became apparent at higher concentrations and with prolonged exposure. The photosynthetic efficiency of C. vulgaris recovered, in contrast to that of M. aeruginosa, which exhibited limited recovery. Spinetoram more significantly inhibited the effective quantum yield of PSII (EQY) in M. aeruginosa than in C. vulgaris. Although spinetoram is not designed to target phytoplankton, its toxicity can disrupt primary productivity and modify phytoplankton-consumer interactions via bottom-up control mechanisms. This study enhances our understanding of spinetoram's ecotoxicity and potential effects on aquatic ecosystems.

Tri-layered core-shell structured deferoxamine magnetic particles promote Microcystis aeruginosa growth.

This experiment prepared magnetic composite siderophores (DMPs) with strong magnetism, excellent adsorption capacity, and high specific surface area. Exploring the synergistic effect of magnetic nanoparticles and siderophores on Microcystis aeruginosa growth under iron-deficient condition, by utilizing the characteristics of the three-layer core-shell structure of DMPs. This study elucidated the potential mechanism by which DMPs promote the cyanobacterial growth through physiological indicators and transcriptome analysis. On the experiment's final day, cell density in DMPs treatment group at 2, 4, and 8 mg/L were 1.10, 1.14 and 1.16 times higher than those in the control group (Ct), respectively. Similarly, chlorophyll and photosynthetic efficiency results showed improved algae growth with increasing DMPs dosage. The microcystin content in DMPs experimental groups at low, medium, and high concentration were 0.91, 0.86, and 0.83 times that of Ct, indicating alleviation of iron deficiency stress. Additionally, based on extracellular polymers, intracellular and extracellular siderophores, and visualization techniques, DMPs nanoparticles captured free iron sources in the environment, promoting algae growth by entering algal cells and facilitating the uptake and utilization of free iron ions from the solution. During the experiment, the iron uptake and transport genes (feoA and feoB) were significantly upregulated, whereas the algal siderophore synthesis gene (pchF) and the TonB-dependent transport system gene (TonB_C) were significantly downregulated, suggesting heightened activity in intracellular iron uptake and transport. This indicates an abundance of intracellular iron, eliminating the need for secrete siderophores to overcome iron deficiency. Microcystis aeruginosa increased iron bioavailability by using iron transported through DMPs in the environment while internalizing these DMPs. This study explored the mechanism of this synergistic effect to boost algal growth, and provided new ideas for elucidating the mechanism of cyanobacterial bloom outbreaks as well as the innovative application of biotechnology.

Identification of algicidal monoterpenoids from four chemotypes of Cinnamomum camphora and their algicidal mechanisms on Microcystis aeruginosa.

Cyanobacterial blooms cause serious environmental issues, and plant secondary metabolites are considered as new algaecide for controlling them. Cinnamomum camphora produces a wide spectrum of terpenoids and has 4 main chemotypes, including linalool, camphor, eucalyptol and borneol chemotype. To develop the new cyanobacterial algaecide by using suitable chemotype of Cinnamomum camphora and the main terpenoids, we analyzed the terpenoid composition in the 4 chemotype extracts, evaluated the algicidal effects of the extracts and their typical monoterpenoids on Microcystis aeruginosa, and investigated the algicidal mechanism of the stronger algicidal agents. Among the 4 chemotypes, eucalyptol and borneol chemotype extracts exhibited stronger algicidal effects. In the 4 chemotype extracts, monoterpenoids were the main compounds, of which linalool, camphor, eucalyptol and borneol were the typical components. Among the 4 typical monoterpenoids, eucalyptol and borneol showed stronger algicidal effects, which killed 78.8% and 100% M. aeruginosa cells, respectively, at 1.2 mM after 48 h. In 1.2 mM eucalyptol and borneol treatments, the reactive oxygen species levels markedly increased, and the caspase-3-like activity also raised. With prolonging the treatment time, M. aeruginosa cells gradually shrank and wrinkled, and the cell TUNEL fluorescence intensity and DNA degradation gradually enhanced, indicating that the lethal mechanism is causing apoptosis-like programmed cell death (PCD). Therefore, eucalyptol and borneol chemotype extracts and their typical monoterpenoids have the potential for developing as algaecides to control cyanobacteria through triggering apoptosis-like PCD.

Sphingomonas lacusdianchii sp. nov., an attached bacterium inhibited by metabolites from its symbiotic cyanobacterium.

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 C18:1ω7c and C16:0 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).

Biotechnological approaches for suppressing Microcystis blooms: insights and challenges.

Cyanobacterial harmful algal blooms, particularly those dominated by Microcystis, pose significant ecological and health risks worldwide. This review provides an overview of the latest advances in biotechnological approaches for mitigating Microcystis blooms, focusing on cyanobactericidal bacteria, fungi, eukaryotic microalgae, zooplankton, aquatic plants, and cyanophages. Recently, promising results have been obtained using cyanobactericidal bacteria: not through the inoculation of cultured bacteria, but rather by nurturing those already present in the periphyton or biofilms of aquatic plants. Fungi and eukaryotic microalgae also exhibit algicidal properties; however, their practical applications still face challenges. Zooplankton grazing on Microcystis can improve water quality, but hurdles exist because of the colonial form and toxin production of Microcystis. Aquatic plants control blooms through allelopathy and nutrient absorption. Although cyanophages hold promise for Microcystis control, their strain-specificity hinders widespread use. Despite successful laboratory validation, field applications of biological methods are limited. Future research should leverage advanced molecular and bioinformatic techniques to understand microbial interactions during blooms and offer insights into innovative control strategies. Despite progress, the efficacy of biological methods under field conditions requires further verification, emphasizing the importance of integrating advanced multi-meta-omics techniques with practical applications to address the challenges posed by Microcystis blooms. KEY POINTS: • A diverse range of biotechnological methods is presented for suppressing Microcystis blooms. • Efficacy in laboratory experiments needs to be proved further in field applications. • Multi-meta-omics techniques offer novel insights into Microcystis dynamics and interactions.

Response of particle-attached and free-living bacterial communities to Microcystis blooms.

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.