Cyanopeptides occurrence and diversity in a Brazilian tropical reservoir: Exploring relationships with water quality.

Microcystins (MCs) are a class of toxic secondary metabolites produced by some cyanobacteria strains that endanger aquatic and terrestrial organisms in various freshwater systems. Although patterns in MC occurrence are being recognized, divergences in the global data still hamper our ability to predict the toxicity of cyanobacterial blooms. This study aimed (i) to determine the dynamics of MCs and other cyanopeptides in a tropical reservoir, (ii) to investigate the correlation between peptides and potential cyanotoxin producers (iii) identifying the possible abiotic factors that influence the peptides. We analyzed, monthly, eight MC variants (MC-RR, -LA, -LF, -LR, -LW, -YR, [D-Asp3]-RR and [D-Asp3]-LR) and other peptides in 47 water samples collected monthly, all season long, from two sampling sites in a tropical eutrophic freshwater reservoir, in southeastern Brazil. The cyanopeptides were assessed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The biomass of potential cyanobacterial producers and water quality variables were measured. MCs were detected in both sampling sites year-round; the total MC concentration varied from 0.21 to 4.04 µg L-1, and three MC variants were identified and quantified (MC-RR, [D-Asp3]-RR, -LR). Additionally, we identified 28 compounds belonging to three other cyanopeptide classes: aeruginosin, microginin, and cyanopeptolin. As potential MC producers, Microcystis spp. and Dolichospermum circinalis were dominant during the study, representing up to 75% of the total phytoplankton. Correlational and redundancy analysis suggested positive effects of dissolved oxygen, nitrate, and total phosphorus on MC and microginins concentration, while water temperature appeared to favor aeruginosins. A comparison between our results and historical data showed a reduction in total phosphorus and cyanobacteria, suggesting increased water quality in the reservoir. However, the current MC concentrations indicate a rise in cyanobacterial toxicity over the last eight years. Moreover, our study underscores the pressing need to explore cyanopeptides other than MCs in tropical aquatic systems.

Cyanotoxins and water quality parameters as risk assessment indicators for aquatic life in reservoirs.

We assessed the extent of pollution in an essential public water supply reservoir (southeastern Brazil). An environmental monitoring study was performed at the Billings Reservoir (at the water catchment site) to assess the water quality in 2017, 2018, and 2019. Physicochemical parameters were analyzed, quantifying the total cyanobacteria and the cyanotoxins microcystins (MCs) and saxitoxins (SXTs), as well as their possible ecological risk to the aquatic environment. We also determined metals and metalloids (As, Ba, Cd, Pb, Cu, Cr, Fe, Mn, Ni, Zn, and Sb) and fecal bacteria (Escherichia coli). Monthly samplings were performed for 2017, 2018, and 2019 (totaling 36 sampling campaigns). Metals, metalloids, and E. coli values were below the maximum limit allowed by the Brazilian legislation. High concentrations of total cyanobacteria (3.07 × 104 - 3.23 × 105 cells/mL), microcystin variants MC-LR (0.67-23.63 µg/L), MC-LA (0.03-8.66 µg/L), MC-RR (0.56-7.92 µg/L), and MC-YR (0.04-1.24 µg/L), as well as the saxitoxins GTX2 (0.18-5.37 µg/L), GTX3 (0.13-4.40 µg/L), and STX (0.12-2.92 µg/L) were detected. From an ecotoxicological point of view, the estimated values for the risk quotient (RQ) for microcystins and saxitoxins were largely greater than 1, indicating a high risk to aquatic life. Therefore, further efforts need to be made to delay the eutrophication of the reservoir.

Temporal and spatial variation in the efficiency of a Floc & Sink technique for controlling cyanobacterial blooms in a tropical reservoir.

One of the main symptoms of eutrophication is the proliferation of phytoplankton biomass, including nuisance cyanobacteria. Reduction of the external nutrient load is essential to control eutrophication, and in-lake interventions are suggested for mitigating cyanobacterial blooms to accelerate ecosystem recovery. Floc & Sink (F&S) is one such intervention technique that consists of applying a low dose of coagulants in combination with ballasts for removing cyanobacteria biomass. It is especially suitable for deep lakes with an external nutrient load that is higher than the internal load and suffers from perennial cyanobacterial bloom events. Studies showing the efficacy of the F&S technique have been published, but those testing its variation in efficacy with changes in the environmental conditions are still scarce. Therefore, we evaluated the efficiency of the F&S technique to remove cyanobacteria from water samples collected monthly from two different sites in a deep tropical reservoir (Funil Reservoir, Brazil) in the laboratory. We tested the efficacy of two coagulants, chitosan (CHI) and poly-aluminum chloride (PAC), alone and in combination with lanthanum-modified bentonite (LMB) in settling phytoplankton biomass. We hypothesized that: ⅰ) the combined treatments are more effective in removing the algal biomass and ⅱ) the efficiency of F&S treatments varies spatially and monthly due to changes in environmental conditions. The combined treatments (PAC + LMB or CHI + LMB) removed up to seven times more biomass than single treatments (PAC, CHI, or LMB). Only the treatments CHI and LMB + CHI differed in efficiency between the sites, although all treatments showed significant variation in efficiency over the months at both the sampling sites. The combined treatments exhibited lower removal efficacy during the warm-rainy months (October-March) than during the mild-cold dry months (April-September). At high pH (pH > 10), the efficiency of the CHI and LMB + CHI treatments decreased. CHI had lower removal efficiency when single-cell cyanobacteria were abundant, while the combined treatments were equally efficient regardless of the morphology of the cyanobacteria. Hence, the combination of PAC as a coagulant with a ballast LMB is the most effective technique to precipitate cyanobacteria under the conditions that are encountered around the year in this tropical reservoir.

'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).

Increasing Temperature Counteracts the Negative Effect of UV Radiation on Growth and Photosynthetic Efficiency of Microcystis aeruginosa and Raphidiopsis raciborskii.

High temperature can promote cyanobacterial blooms, whereas ultraviolet radiation (UVR) can potentially depress cyanobacterial growth by damaging their photosynthetic apparatus. Although the damaging effect of UVR has been well documented, reports on the interactive effects of UV radiation exposure and warming on cyanobacteria remain scarce. To better understand the combined effects of temperature and UVR on cyanobacteria, two strains of nuisance species, Microcystis aeruginosa (MIRF) and Raphidiopsis raciborskii (formerly Cylindrospermopsis raciborskii, CYRF), were grown at 24°C and 28°C and were daily exposed to UVA + UVB (PAR + UVA+UVB) or only UVA (PAR + UVA) radiation. MIRF and CYRF growth rates were most affected by PAR + UVA+UVB treatment and to a lesser extent by the PAR + UVA treatment. Negative UVR effects on growth, Photosystem II (PSII) efficiency and photosynthesis were pronounced at 24°C when compared to that at 28°C. Our results showed a cumulative negative effect on PSII efficiency in MIRF, but not in CYRF. Hence, although higher temperature ameliorates UVR damage, interspecific differences may lead to deviating impacts on different species, and combined elevated temperature and UVR stress could influence species competition.

Downstream transport processes modulate the effects of environmental heterogeneity on riverine phytoplankton.

Environmental heterogeneity (EH) in space and time promotes niche-partition, which leads to high variation in biological communities, such as in algae. In streams, EH is highly related to the intensity of the water flow and may lead to community variation mainly during the low flow conditions. Despite the wide knowledge on the responses of phytoplankton communities to EH in lentic and semi-lentic systems, studies of riverine phytoplankton community variation are still scarce. Here, we first investigated the relationship between phytoplankton community variation and EH in different courses of the river and between seasons. We expected that under low or intermediate flow conditions, there is a positive correlation between community variation and EH. Alternatively, we did not expect any relationship between EH and community variation under high flow condition because stronger downstream transport would mask environmental filtering. We sampled nine sites monthly (May 2012 to April 2013) in a tropical river of Brazilian Southeast. We calculated EH from abiotic data whereas for community variation, here community distinctiveness (CD), we used Sorensen (CDSor) and Bray-Curtis (CDBray) dissimilarities. Differences in EH, CDSor and CDBray were tested at between-season and among-course levels. We found lower distinctiveness during the dry season when EH was the highest. Contrastingly, phytoplankton CD was the highest even when EH was low during the wet season. We found that this pattern raised from the increasing in individuals dispersal during the wet season, promoting mass effects. Finally, our results thus reject the first hypothesis and show a negative relationship between EH and distinctiveness. However, results support our alternative hypothesis and show that during the wet season, distinctiveness is not driven by EH. These results provide new insights into how EH drives community variation, being useful for both basic research about riverine algal communities and biomonitoring programs using phytoplankton communities as bioindicators.

Intraspecific variability in response to phosphorus depleted conditions in the cyanobacteria Microcystis aeruginosa and Raphidiopsis raciborskii.

Phosphorus loading plays an important role in the occurrence of cyanobacterial blooms and understanding how this nutrient affects the physiology of cyanobacteria is imperative to manage these phenomena. Microcystis aeruginosa and Raphidiopsis raciborskii are cyanobacterial species that form potentially toxic blooms in freshwater ecosystems worldwide. Blooms comprise numerous strains with high trait variability, which can contribute to the widespread distribution of these species. Here, we explored the intraspecific variability in response to phosphorus depleted conditions (P-) testing five strains of each species. Strains could be differentiated by cell volume or genetic profiles except for those of the same species, sampling location and date, though these presented differences in their response to (P-). Although differently affected by (P-) over 10 days, all strains were able to grow and maintain photosynthetic activity. For most M. aeruginosa and R. raciborskii strains growth rates were not significantly different comparing (P+) and (P-) conditions. After ten days in (P-), only one M. aeruginosa strain and two R. raciborskii strains showed reduction in biovolume yield as compared to (P+) but in most strains chlorophyll-a concentrations were lower in (P-) than in (P+). Reduced photosystem II efficiency was found for only one R. raciborskii strain while all M. aeruginosa strains were affected. Only two M. aeruginosa and one R. raciborskii strain increased alkaline phosphatase activity under (P-) as compared to (P+). Variation in P-uptake was also observed but comparison among strains yielded homogeneous groups comprised of representatives of both species. Comparing the response of each species as a whole, the (P-) condition affected growth rate, biovolume yield and chlorophyll yield. However, these parameters revealed variation among strains of the same species to the extent that differences between M. aeruginosa and R. raciborskii were not significant. Taken together, these results do not support the idea that R. raciborskii, as a species, can withstand phosphorus limitation better than M. aeruginosa and also point that the level of intraspecific variation may preclude generalizations based on studies that use only one or few strains.

Assessment of possible solid-phase phosphate sorbents to mitigate eutrophication: Influence of pH and anoxia.

Managing eutrophication remains a challenge to water managers. Currently, the manipulation of biogeochemical processes (i.e., geo-engineering) by using phosphorus-adsorptive techniques has been recognized as an appropriate tool to manage the problem. The first step in finding potential mitigating materials is conducting a sequence of upscaling studies that commence with controlled laboratory experiments. Here, the abilities of 10 possible solid-phase-sorbents (SPS) to adsorb P were examined. Four materials adsorbed P, and two of these materials were modified, i.e., a lanthanum-modified-bentonite (LMB) and an aluminum-modified-zeolite (AMZ), and had the highest adsorption capacities of 11.4 and 8.9mgPg-1, respectively. Two natural materials, a red soil (RS) and a bauxite (BAU), were less efficient with adsorption capacities of 2.9 and 3.4mgPg-1, respectively. Elemental composition was not related to P adsorption. Since SPS might be affected by pH and redox status, we also tested these materials at pH values of 6, 7, 8 and 9 and under anoxic condition. All tested materials experienced decreased adsorption capacities under anoxic condition, with maximum adsorptions of 5.3mgPg-1 for LMB, 5.9mgPg-1 for AMZ, 0.2mgPg-1 for RS and 0.2mgPg-1 for BAU. All materials were able to adsorb P across the range of pH values that were tested. The maximum adsorption capacities of LMB and RS were highest at pH6, AMZ was higher at a pH of 9 and BAU at a pH of 8. Thus, pH influenced P adsorption differently. Given the effects of pH and anoxia, other abiotic variables should also be considered. Considering the criteria that classify a useful SPS (i.e., effective, easy to produce, cheap and safe), only the two modified materials that were tested seem to be suitable for upscaling to enclosure studies with anoxic sediments.

Critical assessment of chitosan as coagulant to remove cyanobacteria.

Removal of cyanobacteria from the water column using a coagulant and a ballast compound is a promising technique to mitigate nuisance. As coagulant the organic, biodegradable polymer chitosan has been promoted. Results in this study show that elevated pH, as may be common during cyanobacterial blooms, as well as high alkalinity may hamper the coagulation of chitosan and thus impair its ability to effectively remove positively buoyant cyanobacteria from the water column. The underlying mechanism is likely a shielding of the protonated groups by anions. Inasmuch as there are many chitosan formulations, thorough testing of each chitosan prior to its application is essential. Results obtained in glass tubes were similar to those from standard jar tests demonstrating that glass tube tests can be used for testing effects of coagulants and ballasts in cyanobacteria removal whilst allowing far more replicates. There was no relation between zeta potential and precipitated cyanobacteria. Given the well-known antibacterial activity of chitosan and recent findings of anti-cyanobacterial effects, pre-application tests are needed to decipher if chitosan may cause cell leakage of cyanotoxins. Efficiency- and side-effect testing are crucial for water managers to determine if the selected approach can be used in tailor-made interventions to control cyanobacterial blooms and to mitigate eutrophication.

Chitosan as coagulant on cyanobacteria in lake restoration management may cause rapid cell lysis.

Combining coagulant and ballast to remove cyanobacteria from the water column is a promising restoration technique to mitigate cyanobacterial nuisance in surface waters. The organic, biodegradable polymer chitosan has been promoted as a coagulant and is viewed as non-toxic. In this study, we show that chitosan may rapidly compromise membrane integrity and kill certain cyanobacteria leading to release of cell contents in the water. A strain of Cylindrospermopsis raciborskii and one strain of Planktothrix agardhii were most sensitive. A 1.3 h exposure to a low dose of 0.5 mg l-1 chitosan already almost completely killed these cultures resulting in release of cell contents. After 24 h, reductions in PSII efficiencies of all cyanobacteria tested were observed. EC50 values varied from around 0.5 mg l-1 chitosan for the two sensitive strains, via about 5 mg l-1 chitosan for an Aphanizomenon flos-aquae strain, a toxic P. agardhii strain and two Anabaena cylindrica cultures, to more than 8 mg l-1 chitosan for a Microcystis aeruginosa strain and another A. flos-aquae strain. Differences in sensitivity to chitosan might be related to polymeric substances that surround cyanobacteria. Rapid lysis of toxic strains is likely and when chitosan flocking and sinking of cyanobacteria is considered in lake restoration, flocculation efficacy studies should be complemented with investigation on the effects of chitosan on the cyanobacteria assemblage being targeted.

Controlling cyanobacterial blooms through effective flocculation and sedimentation with combined use of flocculants and phosphorus adsorbing natural soil and modified clay.

Eutrophication often results in blooms of toxic cyanobacteria that hamper the use of lakes and reservoirs. In this paper, we experimentally evaluated the efficacy of a metal salt (poly-aluminium chloride, PAC) and chitosan, alone and combined with different doses of the lanthanum modified bentonite Phoslock(®) (LMB) or local red soil (LRS) to sediment positively buoyant cyanobacteria from Funil Reservoir, Brazil, (22°30'S, 44°45'W). We also tested the effect of calcium peroxide (CaO2) on suspended and settled cyanobacterial photosystem efficiency, and evaluated the soluble reactive P (SRP) adsorbing capacity of both LMB and LRS under oxic and anoxic conditions. Our data showed that buoyant cyanobacteria could be flocked and effectively precipitated using a combination of PAC or chitosan with LMB or LRS. The SRP sorption capacity of LMB was higher than that of LRS. The maximum P adsorption was lowered under anoxic conditions especially for LRS ballast. CaO2 addition impaired photosystem efficiency at 1 mg L(-1) or higher and killed precipitated cyanobacteria at 4 mg L(-1) or higher. A drawback was that oxygen production from the peroxide gave positive buoyancy again to the settled flocs. Therefore, further experimentations with slow release pellets are recommended.

Assessment of the Effects of Light Availability on Growth and Competition Between Strains of Planktothrix agardhii and Microcystis aeruginosa.

In this study, we tested the hypothesis that Planktothrix agardhii strains isolated from a tropical water body were better competitors for light than Microcystis aeruginosa strains. These cyanobacteria are common in eutrophic systems, where light is one of the main drivers of phytoplankton, and Planktothrix is considered more shade-adapted and Microcystis more high-light tolerant. First, the effect of light intensities on growth was studied in batch cultures. Next, the minimum requirement of light (I*) and the effect of light limitation on the outcome of competition was investigated in chemostats. All strains showed similar growth at 10 µmol photons m(-2) s(-1), demonstrating the ability of the two species to grow in low light. The optimum light intensity was lower for P. agardhii, but at the highest light intensity, Microcystis strains reached higher biovolume, confirming that P. agardhii has higher sensitivity to high light. Nonetheless, P. agardhii grew in light intensities considered high (500 µmol photons m(-2) s(-1)) for this species. M. aeruginosa showed a higher carrying capacity in light-limited condition, but I* was similar between all the strains. Under light competition, Microcystis strains displaced P. agardhii and dominated. In two cases, there was competitive exclusion and in the other two P. agardhii managed to remain in the system with a low biovolume (≈15%). Our findings not only show that strains of P. agardhii can grow under higher light intensities than generally assumed but also that strains of M. aeruginosa are better competitors for light than supposed. These results help to understand the co-occurrence of these species in tropical environments and the dominance of M. aeruginosa even in low-light conditions.

Light and phosphate competition between Cylindrospermopsis raciborskii and Microcystis aeruginosa is strain dependent.

The hypothesis that outcomes of phosphorus and light competition between Cylindrospermopsis raciborskii and Microcystis aeruginosa are strain dependent was tested experimentally. Critical requirements of phosphorus (P*) and of light (I*) of two strains of each species were determined through monoculture experiments, which indicated a trade-off between species and also between Microcystis strains. Competition experiments between species were performed using the weakest predicted competitors (with the highest values of P* and of I*) and with the strongest predicted competitors (with the lowest values of P* and of I*). Under light limitation, competition between the weakest competitors led C. raciborskii to dominate. Between the strongest competitors, the opposite was observed, M. aeruginosa displaced C. raciborskii, but both strains co-existed in equilibrium. Under phosphate limitation, competition between the weakest competitors led C. raciborskii to exclude M. aeruginosa, and between the strongest competitors, the opposite was observed, M. aeruginosa displaced C. raciborskii, but the system did not reach an equilibrium and both strains were washed out. Hence, outcomes of the competition depended on the pair of competing strains and not only on species or on type of limitation. We concluded that existence of different trade-offs among strains and between species underlie our results showing that C. raciborskii can either dominate or be displaced by M. aeruginosa when exposed to different conditions of light or phosphate limitation.

Estrutura da comunidade fitoplanctônica da lagoa de Juturnaíba, Araruama, RJ, Brasil: uma comparaçäo entre o centro da regiäo limnética, tributários e canal de drenagem / Structure of phytoplankton community of lagoa de Juturnaíba, Araruama, RJ, Brazil: a comparison between centrum of limnetical region, tributaries and drainage channel

Foi comparada a estrutura da comunidade fitoplanctônica da regiäo limnética da Lagoa, com seus tributários e seu canal de drenagem. A composiçäo florística, diversidade, equitabilidade, abundância e análise qualitativa da afinidade, mostraram maiores semelhanças entre o centro da Lagoa e seu canal de drenagem, sendo estes diferenciados dos tributários. A análise quantitativa da afinidade separou os ambientes em funçäo das concentraçöes das populaçöes de algas, e evidenciou a formaçäo de grupos de espécies abundantes, onde destacam-se o grupo de espécies de Oscillatoria, exclusivo das estaçöes lóticas, e o grupo de espécies que apresentam uma maior adaptaçäo às variaçöes ambientais