Physiological and toxicological response of Microcystis aeruginosa BCCUSP232 exposed to Salvinia auriculata extracts.

Microcystis aeruginosa is one of the most predominant freshwater bloom-forming cyanobacterium found globally which is capable of producing toxic secondary metabolites including microcystins that might intoxicate animals and humans when contaminated water or food is ingested. Salvinia auriculata Aubl is one of the plants that might possess bioactive compounds capable of controlling growth and reproduction of M. aeruginosa. The present study aimed to determine the presence of bioactive compounds in S. auriculata extracts and determine alterations occurred in growth and reproduction of M. aeruginosa when exposed to these plant extracts. In addition, this investigation aimed to examine the influence of S. auriculata on antioxidant enzymes detected in M. aeruginosa. The results obtained demonstrated that the aqueous and ethanolic extracts of S. auriculata presented potential for control of cyanobacteria populations, exhibiting algicidal action on M. aeruginosa as well as interfering in antioxidant enzymes activities and parameters associated with oxidative stress. Phytochemical analyses demonstrated the presence of polyphenols and flavonoids content in both extracts. In addition, application of S. auriculata extracts did not produce cytogenotoxicity and/or mutagenicity utilizing Allium cepa test. Therefore, further studies are needed in order to identify and characterize the compounds responsible for these effects on M. aeruginosa and provide information regarding the possible application of S. auriculata in the treatment of drinking water.

The Microcystis-microbiome interactions: origins of the colonial lifestyle.

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.

In Silico Analysis of a GH3 ß-Glucosidase from Microcystis aeruginosa CACIAM 03.

Cyanobacteria are rich sources of secondary metabolites and have the potential to be excellent industrial enzyme producers. ß-glucosidases are extensively employed in processing biomass degradation as they mediate the most crucial step of bioconversion of cellobiose (CBI), hence controlling the efficiency and global rate of biomass hydrolysis. However, the production and availability of these enzymes derived from cyanobacteria remains limited. In this study, we evaluated the ß-glucosidase from Microcystis aeruginosa CACIAM 03 (MaBgl3) and its potential for bioconversion of cellulosic biomass by analyzing primary/secondary structures, predicting physicochemical properties, homology modeling, molecular docking, and simulations of molecular dynamics (MD). The results showed that MaBgl3 derives from an N-terminal domain folded as a distorted ß-barrel, which contains the conserved His-Asp catalytic dyad often found in glycosylases of the GH3 family. The molecular docking results showed relevant interactions with Asp81, Ala271 and Arg444 residues that contribute to the binding process during MD simulation. Moreover, the MD simulation of the MaBgl3 was stable, shown by analyzing the root mean square deviation (RMSD) values and observing favorable binding free energy in both complexes. In addition, experimental data suggest that MaBgl3 could be a potential enzyme for cellobiose-hydrolyzing degradation.

Effect of hydrological modification on the potential toxicity of Microcystis aeruginosa complex in Salto Grande reservoir, Uruguay.

It is widely known that the environmental conditions caused by the construction of reservoirs favor the proliferation of toxic cyanobacteria and the formation of blooms due to the high residence time of the water, low turbidity, temperature regimes, among others. Microcystin-producing cyanobacteria such as those from the Microcystis aeruginosa complex (MAC) are the most frequently found organisms in reservoirs worldwide, being the role of the environment on microcystin production poorly understood. Here, we addressed the community dynamics and potential toxicity of MAC cyanobacteria in a subtropical reservoir (Salto Grande) located in the low Uruguay river. Samples were taken from five different sites (upstream, inside the reservoir and downstream) during contrasting seasons (summer and winter) to analyze: (i) the MAC community structure by amplicon sequencing of the phycocyanin gene spacer, (ii) the genotype diversity of microcystin-producing MAC by high resolution melting analysis of the mcyJ gene, and (iii) the abundance and mcy transcription activity of the microcystin-producing (toxic) fraction. We found that MAC diversity decreased from summer to winter but, despite the observed changes in MAC community structure, the abundance of toxic organisms and the transcription of mcy genes were always higher inside the reservoir, regardless of the season. Two different genotypes of toxic MAC were detected inside the reservoir, one associated with low water temperature (15 °C) and one thriving at high water temperature (31 °C). These findings indicate that the environmental conditions inside the reservoir reduce community diversity while promoting the proliferation of toxic genotypes that actively transcribe mcy genes, whose relative abundance will depend on the water temperature.

Effect of three commercial algaecides on cyanobacteria and microcystin-LR: implications for drinking water treatment using activated carbon.

Toxic cyanobacterial blooms in aquatic ecosystems are associated to both public health and environmental concerns worldwide. Depending on the treatment technologies used, the removal capacity of cyanotoxins by drinking water treatment plants (DWTPs) is not sufficient to reach safe levels in drinking water. Likewise, controlling these blooms with algaecide may impair the efficiency of DWTPs due to the possible lysis of cyanobacterial cells and consequent release of cyanotoxins. We investigated the effects of three commercial algaecides (cationic polymer, copper sulfate, and hydrogen peroxide) on the growth parameters of the cyanobacterium Microcystis aeruginosa and the release of microcystin-LR (MC-LR). The potential interference of each algaecide on the MC-LR removal by adsorption on activated carbon (AC) was also tested through adsorption isotherms and kinetics experiments. Most algaecides significantly decreased the cell density and biovolume of M. aeruginosa, as well as increased the release of MC-LR. Interestingly, the presence of the algaecides in binary mixtures with MC-LR affected the adsorption of the cyanotoxin. Relevant adsorption parameters (e.g., maximum adsorption capacity, adsorption intensity, and affinity between MC-LR and AC) were altered when the algaecides were present, especially in the case of the cationic polymer. Also, the algaecides influenced the kinetics (e.g., by shifting the initial adsorption and the desorption constant), which may directly affect the design and operation of DWTPs. Our study indicated that algaecides can significantly impact the fate and the removal of MC-LR in DWTPs when the adsorption process is employed, with important implications for the management and performance of such facilities.

A Promising Use of Trimethyl Chitosan for Removing Microcystis aeruginosa in Water Treatment Processes.

The increase in cyanobacterial blooms linked to climate change and the eutrophication of water bodies is a global concern. The harmful cyanobacterium Microcystis aeruginosa is one of the most common bloom-forming species whose removal from fresh water and, in particular, from that used for water treatment processes, remains a crucial goal. Different biodegradable and environmentally friendly coagulants/flocculants have been assayed, with chitosan showing a very good performance. However, chitosan in its original form is of limited applicability since it is only soluble in acid solution. The objective of this work was therefore to test the coagulant/flocculant capacity of trimethylchitosan (TMC), a chitosan derivative produced from residues of the fishing industry. TMC has a constitutively net positive charge enabling it to remain in solution regardless of the pH. Results show that even at alkaline pHs, common during cyanobacterial blooms, TMC is effective in removing buoyant cyanobacteria from the water column, both in test tube and Jar-Test experiments. Cell integrity was confirmed by fluorescent stain and electron microscopy. Our findings lead us to conclude that the use of TMC to remove bloom cells early in the treatment of drinking water is both feasible and promising.

Genotyping and Multivariate Regression Trees Reveal Ecological Diversification within the Microcystis aeruginosa Complex along a Wide Environmental Gradient.

Addressing the ecological and evolutionary processes underlying biodiversity patterns is essential to identify the mechanisms shaping community structure and function. In bacteria, the formation of new ecologically distinct populations (ecotypes) is proposed as one of the main drivers of diversification. New ecotypes arise when mutations in key functional genes or acquisition of new metabolic pathways by horizontal gene transfer allow the population to exploit new resources, permitting their coexistence with the parental population. We previously reported the presence of microcystin-producing organisms of the Microcystis aeruginosa complex (toxic MAC) through an 800-km environmental gradient ranging from freshwater to estuarine-marine waters in South America. We hypothesize that the success of toxic MAC in such a gradient is due to the existence of very closely related populations that are ecologically distinct (ecotypes), each specialized to a specific arrangement of environmental variables. Here, we analyzed toxic MAC genetic diversity through quantitative PCR (qPCR) and high-resolution melting analysis (HRMA) of a functional gene (mcyJ, microcystin synthetase cluster). We explored the variability of the mcyJ gene along the environmental gradient by multivariate classification and regression trees (mCART). Six groups of mcyJ genotypes were distinguished and associated with different combinations of water temperature, conductivity, and turbidity. We propose that each mcyJ variant associated with a defined environmental condition is an ecotype (or species) whose relative abundances vary according to their fitness in the local environment. This mechanism would explain the success of toxic MAC in such a wide array of environmental conditions. IMPORTANCE Organisms of the Microcystis aeruginosa complex form harmful algal blooms (HABs) in nutrient-rich water bodies worldwide. MAC HABs are difficult to manage owing to the production of potent toxins (microcystins) that resist water treatment. In addition, the role of microcystins in the ecology of MAC organisms is still elusive, meaning that the environmental conditions driving the toxicity of the bloom are not clear. Furthermore, the lack of coherence between morphology-based and genomic-based species classification makes it difficult to draw sound conclusions about when and where each member species of the MAC will dominate the bloom. Here, we propose that the diversification process and success of toxic MAC in a wide range of water bodies involves the generation of ecotypes, each specialized in a particular niche, whose relative abundance varies according to its fitness in the local environment. This knowledge can improve the generation of accurate prediction models of MAC growth and toxicity, helping to prevent human and animal intoxication.

Microcystins can be extracted from Microcystis aeruginosa using amino acid-derived biosurfactants.

Microcystin, a cyanotoxin produced by Microcystis aeruginosa growing in eutrophic waters, can promote liver tumors in people ingesting contaminated water. To date, water treatment systems have not been effective in removing or degrading these cyanotoxins. In this work, we investigated the inhibitory activity of surfactants on the growth of M. aeruginosa and their application to extract the intracellular produced cyanotoxins. The experiments involving growth inhibition and extraction of cyanotoxins were carried out using the non-biodegradable surfactant cetyl trimethyl ammonium bromide (CTAB) in addition to other biodegradable surfactants. These were Tween 80 and surfactants derived from amino acids and peptides, respectively, from arginine, SDA, and hydrolyzed peptone, SDP. We demonstrated that the tested surfactants could be used to inhibit the growth of M. aeruginosa. At this point, CTAB and SDA proved to be the most competent surfactants in reducing cyanobacterial growth. Moreover, microcystins have been successfully removed from the water employing a cloud point extraction protocol based on the use of these surfactants and ammonium sulfate.

A Stochastic Characterization of Hydrogen Peroxide-Induced Regulated Cell Death in Microcystis aeruginosa.

Regulated cell death (RCD) encompasses the activation of cellular pathways that initiate and execute a self-dismissal process. RCD occur over a range of stressors doses that overcome pro-survival cellular pathways, while higher doses cause excessive damage leading to passive accidental cell death (ACD). Hydrogen peroxide (HP) has been proposed as a potential tool to control harmful cyanobacterial blooms, given its capacity to remove cyanobacterial cells and oxidize cyanotoxins. HP is a source of hydroxyl radicals and is expected to induce RCD only within a limited range of concentrations. This property makes this compound very useful to better understand stress-driven RCD. In this work, we analyzed cell death in microcystin-producing Microcystis aeruginosa by means of a stochastic dose response model using a wide range of HP concentrations (0, 0.29, 1.76, 3.67, 7.35, 14.70, and 29.5 mM). We used flow cytometry and unsupervised classification to study cell viability and characterize transitional cell death phenotypes after exposing cells to HP for 48 and 72 h. Non-linear regression was used to fit experimental data to a logistic cumulative distribution function (cdf) and calculate the half maximal effective concentration (EC50). The EC50 of M. aeruginosa exposed to HP were 3.77 ± 0.26 mM and 4.26 ± 0.22 mM at 48 and 72 h, respectively. The derivative of cdf (probability density function; pdf) provided theoretical and practical demonstration that EC50 is the minimal dose required to cause RCD in 50% of cells, therefore maximizing the probability of RCD occurrence. 1.76 mM HP lead to an antioxidant stress response characterized by increased reactive oxygen species (ROS) levels and HP decomposition activity. The exposure of 3.67 mM HP induced a dose-related transition in cell death phenotype, and produced several morphological changes (a less dense stroma, distortion of the cell membrane, partial disintegration of thylakoids, extensive cytoplasmic vacuolation and highly condensed chromatin). The EC50 and the stochastic cdf and pdf together with the multidimensional transitional phenotypic analysis of single cells contribute to further characterize cell death pathways in cyanobacteria.

'Floc and Sink' Technique Removes Cyanobacteria and Microcystins from Tropical Reservoir Water.

Combining coagulants with ballast (natural soil or modified clay) to remove cyanobacteria from the water column is a promising tool to mitigate nuisance blooms. Nevertheless, the possible effects of this technique on different toxin-producing cyanobacteria species have not been thoroughly investigated. This laboratory study evaluated the potential effects of the "Floc and Sink" technique on releasing microcystins (MC) from the precipitated biomass. A combined treatment of polyaluminium chloride (PAC) with lanthanum modified bentonite (LMB) and/or local red soil (LRS) was applied to the bloom material (mainly Dolichospermum circinalis and Microcystis aeruginosa) of a tropical reservoir. Intra and extracellular MC and biomass removal were evaluated. PAC alone was not efficient to remove the biomass, while PAC + LMB + LRS was the most efficient and removed 4.3-7.5 times more biomass than other treatments. Intracellular MC concentrations ranged between 12 and 2.180 µg L-1 independent from the biomass. PAC treatment increased extracellular MC concentrations from 3.5 to 6 times. However, when combined with ballast, extracellular MC was up to 4.2 times lower in the top of the test tubes. Nevertheless, PAC + LRS and PAC + LMB + LRS treatments showed extracellular MC concentration eight times higher than controls in the bottom. Our results showed that Floc and Sink appears to be more promising in removing cyanobacteria and extracellular MC from the water column than a sole coagulant (PAC).

Physiological responses and specific fatty acids composition of Microcystis aeruginosa exposed to total solar radiation and increased temperature.

The combined effects of increased temperature and solar ultraviolet radiation (UVR, 280-400 nm) on M. aeruginosa cultures was analyzed in terms of cell abundance, reactive oxygen and nitrogen species (ROS/RNS), antioxidant activity of catalase (CAT), superoxide dismutase (SOD), glutathione S transferase (GST), fatty acids (FA) content and lipid damage. After 12 days exposure to high temperature (29 °C), cells were exposed to solar UVR (4 h). Ultraviolet-B radiation (UVBR, 280-315 nm) resulted into low cell abundance, high ROS/RNS and a significant increase in SOD activity with no changes in GST and a decreased CAT activity at control temperature (26 °C). A significant increase in the analyzed enzymatic antioxidants was observed at 29 °C, as a response to avoid ultraviolet-A radiation (UVAR, 315-400 nm) damage. The relative abundance of ω6 FAs was not affected by UVAR while ω3 FA were highly sensitive at 29 °C but unsaturated fatty acids (UFA) peroxidation did not occur. The differential response in FA to high temperature and UVAR results in differences in lipid damage and antioxidants. It was evident that selected UFAs (mostly ω6) play an important role in high temperature adaptation in addition to enzymatic antioxidant increased activity shifting the temperature growth from 26 to 29 °C. Thus, cell death and UFA damage were avoided at high temperature and low solar irradiance thanks to an increased enzymatic antioxidant activity.

Hazardous cyanobacteria integrity response to velocity gradient and powdered activated carbon in water treatment plants.

Although some studies have investigated the impact caused by chemicals used on water treatment (coagulants and oxidants) on cyanobacteria integrity, the isolated effect of shear stress during coagulation is still not fully understood. This study evaluated the impact of different velocity gradients, mixing times, and the addition of powdered activated carbon (PAC) on the integrity of Microcystis aeruginosa, Raphidiopsis raciborskii, and Dolichospermum circinale, known producers of toxin and taste and odor (T&O) compounds. No association was found between R. raciborskii cell lysis and velocity gradient, with or without PAC, demonstrating the high resilience of this taxon to shear stress. In contrast, an association was found for M. aeruginosa at the highest velocity gradient evaluated (1000 s-1) and for D. circinale above the lowest velocity gradient studied (600 s-1). After PAC addition, there was a reduction in the chances of finding M. aeruginosa intact cells above velocity gradient 800 s-1 at 45 s, while D. circinale show cell lysis in all the scenarios expect at 600 s-1 and 10 s of agitation. The additional impact of PAC on cell lysis may lead to more release of metabolites and shows the need to adjust the hydraulic conditions in the rapid mixing stage, especially when more "fragile" cyanobacteria are present. Neither cyanobacterial cell size nor morphology was shown to be relevant to shear stress sensitivity, indicating that cell wall composition might have been an important factor in controlling cell lysis.

Occurrence and diversity of viruses associated with cyanobacterial communities in a Brazilian freshwater reservoir.

As part of the phytoplankton of marine and freshwater environments around the world, cyanobacteria interact with viruses (cyanophages) that affect their abundance and diversity. Investigations focusing on cyanophages co-occurring with freshwater cyanobacteria are scarce, particularly in Brazil. The aim of this study was to assess the diversity of cyanophages associated with a Microcystis-dominated cyanobacterial bloom in a tropical reservoir. Samples were processed as viral fractions of water and cellular fractions, and temporal fluctuations in the abundance of Ma-LMM01-type cyanophages and their Microcystis hosts were determined by qPCR. We applied shotgun metagenomics to obtain a wider characterization of the cyanophage community. During the study period, Microcystis gene copies were quantified in all cellular fractions, and the copy number of the Ma-LMM01 phage gene tended to increase with host abundance. Metagenomic analysis demonstrated that Caudovirales was the major viral order associated with the cyanophage families Myoviridae (34-88%), Podoviridae (3-42%), and Siphoviridae (6-23%). The metagenomic analysis results confirmed the presence of Microcystis cyanophages in both viral and cellular fractions and demonstrated a high relative abundance of picocyanobacteria-related viruses and Prochlorococcus (36-52%) and Synechococcus (37-50%) phages. For other main cyanobacterial genera, no related cyanophages were identified, which was probably due to the scarce representation of cyanophage sequences in databanks. Thus, the studied reservoir hosted a diverse cyanophage community with a remarkable contribution of phages related to picoplanktonic cyanobacteria. These results provide insights that motivate future sequencing efforts to assess cyanophage diversity and recover complete genomes.

Similar toxicity mechanisms between graphene oxide and oxidized multi-walled carbon nanotubes in Microcystis aeruginosa.

In photosynthetic microorganisms, the toxicity of carbon nanomaterials (CNMs) is typically characterized by a decrease in growth, viability, photosynthesis, as well as the induction of oxidative stress. However, it is currently unclear how the shape of the carbon structure in CNMs, such as in the 1-dimensional carbon nanotubes (CNTs) compared to the two-dimensional graphene oxide (GO), affects the way they interact with cells. In this study, the effects of GO and oxidized multi-walled CNTs were compared in the cyanobacterium Microcystis aeruginosa to determine the similarities or differences in how the two CNMs interact with and induce toxicity to cyanobacteria. Using change in Chlorophyll a concentrations, the effective concentrations inducing 50% inhibition (EC50) at 96 h are found to be 11.1 µg/mL and 7.38 µg/mL for GO and CNTs, respectively. The EC50 of the two CNMs were not found to be statistically different. Changes in fluorescein diacetate and 2',7'-dichlorodihydrofluorescein diacetate fluorescence, measured at the EC50 concentrations, suggest a decrease in esterase enzyme activity but no oxidative stress. Scanning and transmission electron microscopy imaging did not show extensive membrane damage in cells exposed to GO or CNTs. Altogether, the decrease in metabolic activity and photosynthetic activity without oxidative stress or membrane damage support the hypothesis that both GO and CNTs induced indirect toxicity through physical mechanisms associated with light shading and cell aggregation. This indirect toxicity explains why the intrinsic differences in shape, size, and surface properties between CNTs and GO did not result in differences in how they induce toxicity to cyanobacteria.

Rapid freshwater discharge on the coastal ocean as a mean of long distance spreading of an unprecedented toxic cyanobacteria bloom.

Cyanobacterial toxic blooms are a worldwide problem. The Río de la Plata (RdlP) basin makes up about one fourth of South America areal surface, second only to the Amazonian. Intensive agro-industrial land use and the construction of dams have led to generalized eutrophication of main tributaries and increased the intensity and duration of cyanobacteria blooms. Here we analyse the evolution of an exceptional bloom at the low RdlP basin and Atlantic coast during the summer of 2019. A large array of biological, genetic, meteorological, oceanographic and satellite data is combined to discuss the driving mechanisms. The bloom covered the whole stripe of the RdlP estuary and the Uruguayan Atlantic coasts (around 500 km) for approximately 4 months. It was caused by the Microcystis aeruginosa complex (MAC), which produces hepatotoxins (microcystin). Extreme precipitation in the upstream regions of Uruguay and Negro rivers' basins caused high water flows and discharges. The evolution of meteorological and oceanographic conditions as well as the similarity of organisms' traits in the affected area suggest that the bloom originated in eutrophic reservoirs at the lower RdlP basin, Salto Grande in the Uruguay river, and Negro river reservoirs. High temperatures and weak Eastern winds prompted the rapid dispersion of the bloom over the freshwater plume along the RdlP northern and Atlantic coasts. The long-distance rapid drift allowed active MAC organisms to inoculate freshwater bodies from the Atlantic basin, impacting environments relevant for biodiversity conservation. Climate projections for the RdlP basin suggest an increase in precipitation and river water flux, which, in conjunction with agriculture intensification and dams' construction, might turn this extraordinary event into an ordinary situation.

The combined effect of clethodim (herbicide) and nitrogen variation on allelopathic interactions between Microcystis aeruginosa and Raphidiopsis raciborskii.

The large-scale use of herbicides deteriorates water quality and threatens aquatic biodiversity. Unfortunately, there are few studies on the ecological effects of herbicides on toxin-producing strains of cyanobacteria under changing nutrient conditions. The objective of the present study was to investigate the effects of the herbicide clethodim and nitrogen variation on the allelopathic interactions and toxin production of Microcystis aeruginosa BCCUSP232 and Raphidiopsis raciborskii (formerly known as Cylindrospermopsis raciborskii) ITEPA1. M. aeruginosa had increased cell density when exposed to the clethodim (H +) (23.55 mg/L), whereas the highest cell density of R. raciborskii was observed in the treatment with clethodim plus limited nitrogen. Also, the cell-free exudate of R. raciborskii significantly stimulated the growth of M. aeruginosa on day 3 of the experiment. The concentration of chlorophyll-a in M. aeruginosa cultures generally increased in all the treatments, while in R. raciborskii cultures, the opposite occurred. Total microcystins (MCs) content of M. aeruginosa in the mixed cultures was 68% higher in nitrogen-enriched conditions than the control. A similar increase in MC content occurred in M. aeruginosa unialgal culture treated with R. raciborskii exudate. Total saxitoxin concentration was 81% higher in mixed cultures of R. raciborskii simultaneously exposed to high nitrogen and clethodim. Similarly, unialgal cultures of R. raciborskii exposed to either high nitrogen or clethodim had higher saxitoxins concentrations than the control. The intracellular H2O2 content of M. aeruginosa cultures decreased, whereas, in R. raciborskii cultures, it increased during exposure to high nitrogen and clethodim. Only R. raciborskii had a significant variation in peroxidase activity. The activities of glutathione S-transferase of both strains were higher in the presence of clethodim. These results revealed that nitrogen enrichment and the presence of clethodim might lead to the excessive proliferation of M. aeruginosa and R. raciborskii and increased production of cyanotoxins in aquatic environments.

Growth of Microcystisstrains isolated from environments with the presence and absence of submerged macrophytes in coexistence with Ceratophyllum demersum

Cyanobacterial blooms can cause severe ecological and health problems in drinking water reservoirs. To alleviate this problem, allelopathically active submerged macrophytes can be used to reduce cyanobacterial growth. Accordingly, this study aimed to evaluate the sensitivity of strains of the Microcystis aeruginosacomplex isolated from reservoirs with the presence and absence of submerged macrophytes to the allelochemicals of Ceratophyllum demersum.A coexistence experiment was carried out between the submerged macrophyte C. demersum and four Microcystisstrains, with two treatments for each strain, one in coexistence with the submerged macrophyte (7 g L-1) and control (in the absence of the macrophyte). Two strains of M. aeruginosa(BMIUFRPE-06 and BMIUFRPE-07) and two of M. panniformis(BMIUFRPE-08 and BMIUFRPE-09) were used, which were isolated from Cajueiro (with submerged macrophytes) and Tapacurá (without submerged macrophytes) reservoirs, respectively. The biomass of Microcystisstrains from the reservoir without macrophytes (BMIUFRPE-08 and BMIUFRPE-09) was significantly inhibited in 96% (T-test: p 0.05; growth rate -ANOVA: p > 0.05). These results suggest that strains isolated from environments with submerged macrophytes are less sensitive to allelochemicals of these plants,as these strains may be adapted to the coexistence with submerged macrophytes.

Growth of Microcystisstrains isolated from environments with the presence and absence of submerged macrophytes in coexistence with Ceratophyllum demersum

Cyanobacterial blooms can cause severe ecological and health problems in drinking water reservoirs. To alleviate this problem, allelopathically active submerged macrophytes can be used to reduce cyanobacterial growth. Accordingly, this study aimed to evaluate the sensitivity of strains of the Microcystis aeruginosacomplex isolated from reservoirs with the presence and absence of submerged macrophytes to the allelochemicals of Ceratophyllum demersum.A coexistence experiment was carried out between the submerged macrophyte C. demersum and four Microcystisstrains, with two treatments for each strain, one in coexistence with the submerged macrophyte (7 g L-1) and control (in the absence of the macrophyte). Two strains of M. aeruginosa(BMIUFRPE-06 and BMIUFRPE-07) and two of M. panniformis(BMIUFRPE-08 and BMIUFRPE-09) were used, which were isolated from Cajueiro (with submerged macrophytes) and Tapacurá (without submerged macrophytes) reservoirs, respectively. The biomass of Microcystisstrains from the reservoir without macrophytes (BMIUFRPE-08 and BMIUFRPE-09) was significantly inhibited in 96% (T-test: p < 0.01) and 74% (T-test: p< 0.05), when compared to the control, respectively, with lower values of growth rates (ANOVA: p < 0.05). The strains isolated from the reservoir with macrophytes (BMIUFRPE-06 and BMIUFRPE-07) showed a growth delay (biomass reductions of 44 and 58%, respectively) in the coexistence treatment but without significant difference from the control on the sixth day of the experiment (biomass -T-test: p > 0.05; growth rate -ANOVA: p > 0.05). These results suggest that strains isolated from environments with submerged macrophytes are less sensitive to allelochemicals of these plants,as these strains may be adapted to the coexistence with submerged macrophytes.(AU)

Morphology captures toxicity in Microcystis aeruginosa complex: Evidence from a wide environmental gradient✰.

Blooms of the Microcystis aeruginosa complex (MAC) consist of mixtures of toxin-producing and non-toxin-producing populations, but the environmental conditions that determine their relative abundance and shift are not clear. Morphological traits reflect the responses of MAC organisms to environmental changes, thus they could be useful to improve the predictability of the abundance of both toxic and nontoxic populations. In this work, the response of MAC toxic populations to environmental conditions and their relationship with morphology (size of organisms) were investigated in different water bodies (reservoir, river, and estuary) covering wide salinity (0-33) and temperature (10-36 °C) gradients. Sub-surface water samples were collected and divided into 4 size classes (mesh size 〈20 µm, 20-60 µm, 60-150 µm and〉 150 µm) and three toxicity proxies were assessed (mcyE gene and transcripts copy numbers and microcystin concentration) for each size-class. For all the size-classes, the logarithm of the number of mcyE gene copies per sample was proportional to the logarithm of the corresponding biovolume fraction, showing that MAC biovolume is a good indicator of toxicity potential. When toxicity was analyzed through mcyE transcript abundance and microcystin concentration, the largest size fraction (>150 µm) showed the highest toxicity values of both proxies. Nevertheless, mcyE transcription and toxin production per cell were higher in the colonies retained in the 60 to 150 µm size fractions, followed by single cells (<20 µm). At the reservoir, where environmental variability is low, the total abundance of mcyE gene copies was significantly explained by MAC biovolume, regardless of the environmental conditions. However, when data from the reservoir to the estuary were modeled, biovolume and temperature (with a minor contribution of salinity and wind intensity) were selected in the best models. According to these results, the size distribution of MAC biovolume appears as a good predictor of active toxin production, being the colonies in the 60-150 µm size fraction good indicators of higher toxicity. These results can be used to predict MAC toxicity based on the size structure of the community.

Effect of temperature on microcystin-LR removal and lysis activity on Microcystis aeruginosa (cyanobacteria) by an indigenous bacterium belonging to the genus Achromobacter.

Microcystis is a frequent cyanobacterium bloom-forming with cosmopolitan distribution which can produce a hepatotoxin group called microcystins (MCs). These MCs are resistant to the traditional processes employed in the water treatment plants and they are often detected after conventional treatments. Because of this, the bio-removal studies have obtained a great interest in the last decades. In this work, a bacterial strain namely LG1 with the ability to remove microcystin-LR (MC-LR) under laboratory conditions was isolated from Rio de la Plata River and it was identified as Achromobacter spp. This ubiquitous bacterium was able to remove 79.5% MC-LR in 7 days with average removal time of 3.33 ± 0.08, 3.06 ± 0.05, and 2.77 ± 0.05 days at 28, 32, and 36 ± 1 °C, being higher at high temperature (36 °C) with an activation energy = 16.79 ± 1.99 kJ mol-1. LG1 grew better at higher temperature (from 28 to 36 ± 1 °C) increasing the specific growth rate (µ) and reducing 2-fold the lag phase duration (LPD) without significant differences (p > 0.05) between maximum population density (MPD). In addition, LG1 showed a lysis activity on two M. aeruginosa native strains in 7 days measured as chlorophyll a (Chl-a) concentration. The lysis activity increased around 2-fold when increasing the temperature from 28 to 36 ± 1 °C. This is the first report of an indigenous bacterium belonging to the genus Achromobacter spp. isolated from the Rio de la Plata River with the capacity to remove MC-LR and lysis activity on M. aeruginosa.