Synthesis of Cyclopeptides Analogues of Natural Products and Evaluation as Herbicides and Inhibitors of Cyanobacteria.

Natural products derived from plants or microorganisms have been considered as eco-friendly herbicides with application in crop protection. Several natural cyclopeptides have been reported as herbicides, while others have been identified as inhibitors of cyanobacteria. In this work, the syntheses of cyclotetrapeptides and cyclopentapeptides analogues of natural products were successfully performed by solid-phase peptide synthesis of their linear precursor and solution-phase macrolactamization. Four of the obtained peptides and cyclopeptides present phytotoxicity with more than 70% of radicle growth inhibition at 67 µg/mL. In addition, evaluation of 20 peptides and cyclopeptides, as inhibitors of cyanobacteria, rendered five active compounds that reduced the concentration of microcystins in the culture medium.

Cyanobacterial bloom monitoring and assessment in Latin America.

Cyanobacterial blooms have serious adverse effects on human and environmental health. In Latin America, one of the main world's freshwater reserves, information on this phenomenon remains sparse. To assess the current situation, we gathered reports of cyanobacterial blooms and associated cyanotoxins in freshwater bodies from South America and the Caribbean (Latitude 22° N to 45° S) and compiled the regulation and monitoring procedures implemented in each country. As the operational definition of what is a cyanobacterial bloom remains controversial, we also analyzed the criteria used to determine the phenomena in the region. From 2000 to 2019, blooms were reported in 295 water bodies distributed in 14 countries, including shallow and deep lakes, reservoirs, and rivers. Cyanotoxins were found in nine countries and high concentrations of microcystins were reported in all types of water bodies. Blooms were defined according to different, and sometimes arbitrary criteria including qualitative (changes in water color, scum presence), quantitative (abundance), or both. We found 13 different cell abundance thresholds defining bloom events, from 2 × 103 to 1 × 107 cells mL-1. The use of different criteria hampers the estimation of bloom occurrence, and consequently the associated risks and economic impacts. The large differences between countries in terms of number of studies, monitoring efforts, public access to the data and regulations regarding cyanobacteria and cyanotoxins highlights the need to rethink cyanobacterial bloom monitoring, seeking common criteria. General policies leading to solid frameworks based on defined criteria are needed to improve the assessment of cyanobacterial blooms in Latin America. This review represents a starting point toward common approaches for cyanobacterial monitoring and risk assessment, needed to improve regional environmental policies.

Nutrients and not temperature are the key drivers for cyanobacterial biomass in the Americas.

Cyanobacterial blooms imperil the use of freshwater around the globe and present challenges for water management. Studies have suggested that blooms are trigged by high temperatures and nutrient concentrations. While the roles of nitrogen and phosphorus have long been debated, cyanobacterial dominance in phytoplankton has widely been associated with climate warming. However, studies at large geographical scales, covering diverse climate regions and lake depths, are still needed to clarify the drivers of cyanobacterial success. Here, we analyzed data from 464 lakes covering a 14,000 km north-south gradient in the Americas and three lake depth categories. We show that there were no clear trends in cyanobacterial biomass (as biovolume) along latitude or climate gradients, with the exception of lower biomass in polar climates. Phosphorus was the primary resource explaining cyanobacterial biomass in the Americas, while nitrogen was also significant but particularly relevant in very shallow lakes (< 3 m depth). Despite the assessed climatic gradient water temperature was only weakly related to cyanobacterial biomass, suggesting it is overemphasized in current discussions. Depth was critical for predicting cyanobacterial biomass, and shallow lakes proved more vulnerable to eutrophication. Among other variables analyzed, only pH was significantly related to cyanobacteria biomass, likely due to a biologically mediated positive feedback under high nutrient conditions. Solutions toward managing harmful cyanobacteria should thus consider lake morphometric characteristics and emphasize nutrient control, independently of temperature gradients, since local factors are more critical - and more amenable to controls - than global external forces.

Satellite assessment of eutrophication hot spots and algal blooms in small and medium-sized productive reservoirs in Uruguay's main drinking water basin.

Intensive agricultural activities favor eutrophication and harmful phytoplankton blooms due to the high export of nutrients and damming of rivers. Productive watersheds used for water purification can have multiple reservoirs with phytoplankton blooms, which constitutes a high health risk. In general, water quality monitoring does not cover small- and medium-sized reservoirs (0.25-100 ha) of productive use due to their large number and location in private properties. In this work, the in situ trophic state of fourteen reservoirs was simultaneously assessed using Sentinel-2 images in the Santa Lucía River Basin, the main drinking water basin in Uruguay. These reservoirs are hypereutrophic (0.18-5.22 mg total P L-1) with high phytoplankton biomasses (2.8-4439 µg chlorophyll-a L-1), mainly cyanobacteria. Based on data generated in situ and Sentinel-2 imagery, models were fitted to estimate satellite Chl-a and transparency in all the basin reservoirs (n = 486). The best fits were obtained with the green-to-red band ratio (560 and 665 nm, R2 = 0.84) to estimate chlorophyll-a and reflectance at 833 nm (R2 = 0.73) to determine transparency. The spatial distribution of the trophic state was explored by spatial autocorrelation and hotspot analysis, and the variation in spatial patterns could be determined prior and subsequent to a maximum cyanobacteria value in water treatment plant intakes. Therefore, reservoirs with greater potential for phytoplankton biomass export were identified. This work provides the first fitted tool for satellite monitoring of numerous reservoirs and strengthens the country's ability to respond to harmful phytoplankton blooms in its main drinking water basin.

Schindler's legacy: from eutrophic lakes to the phosphorus utilization strategies of cyanobacteria.

David Schindler and his colleagues pioneered studies in the 1970s on the role of phosphorus in stimulating cyanobacterial blooms in North American lakes. Our understanding of the nuances of phosphorus utilization by cyanobacteria has evolved since that time. We review the phosphorus utilization strategies used by cyanobacteria, such as use of organic forms, alternation between passive and active uptake, and luxury storage. While many aspects of physiological responses to phosphorus of cyanobacteria have been measured, our understanding of the critical processes that drive species diversity, adaptation and competition remains limited. We identify persistent critical knowledge gaps, particularly on the adaptation of cyanobacteria to low nutrient concentrations. We propose that traditional discipline-specific studies be adapted and expanded to encompass innovative new methodologies and take advantage of interdisciplinary opportunities among physiologists, molecular biologists, and modellers, to advance our understanding and prediction of toxic cyanobacteria, and ultimately to mitigate the occurrence of blooms.

Nuisance phytoplankton transport is enhanced by high flow in the main river for drinking water in Uruguay.

Eutrophication, climate change, and river flow fragmentation are the main cause of nuisance algal blooms worldwide. This study evaluated the conditions that trigger the growth and occurrence of nuisance phytoplankton in the Santa Lucía River, a subtropical floodplain lotic system that supplies drinking water to 60% of the population of Uruguay. The main variables that explained phytoplankton biovolume were extracted from generalized linear models (GLM). The potential impact of nuisance organism advection on water utility was estimated by the phytoplankton biovolume transport (BVTR, m3 day-1), an indicator of biomass load. Santa Lucía River had a wide flow range (0.7×105-1438×105 m3 day-1) and eutrophic conditions (median, TP: 0.139 mg L-1; TN: 0.589 mg L-1). GLMs indicated that phytoplankton biomass increased with temperature and soluble reactive phosphorus. Contrary to expectations, the presence of cyanobacteria was positively associated with periods of high flow that result in high cyanobacterial biovolume transport, with a probability of 3.35 times higher when flow increased by one standard deviation. The cyanobacterial biovolume transported (max: 9.5 m3 day-1) suggests that biomass was subsidized by allochthonous inocula. Biovolume from other nuisance groups (diatoms, cryptophytes, and euglenophytes) was positively associated with low-flow conditions and high nutrient concentrations in the main river channel, thereby indicating that these conditions boost eukaryote blooms. The evaluation of BVTR allows a better understanding of the dynamics of fluvial phytoplankton and can help to anticipate scenarios of nuisance species transport.

Hotspot analysis of spatial distribution of algae blooms in small and medium water bodies.

Intensive land use favors eutrophication processes and algae bloom proliferation in freshwaters, which is considered to be one of the main environmental issues worldwide. In general, and particularly in South America, inland water monitoring only covers the main water bodies due to the high costs and efforts involved. In order to improve the coverage of spatial and temporal of algae bloom monitoring, remote sensing serves as an alternative tool. Thereby, the analysis of significant spatial clusters of high values (hotspots) and low values (coldspots) of chlorophyll-a has been applied in coastal studies; however, at present, there are no studies in freshwaters. In this study, Getis-Ord Gi* hotspot analysis was applied to detect spatial distribution patterns of algae bloom dynamics in small- and medium-sized freshwater bodies. Four in situ samplings were carried out in five suburban lakes of Uruguay, in agreement with the satellite capture. Total and cyanobacterial chlorophyll-a concentration, and suspended solids were evaluated. Linear models were developed by combining pre-established indexes with additional Sentinel-2 spectral bands and in situ data. The relationship between red and red edge regions allowed mapping the chlorophyll-a in the study lakes with an adjustment of R2 = 0.83. Hotspot analysis was performed with the selected linear model, and significant chlorophyll-a variability within each lake was successfully detected. The novel application of hotspots analyses presented in this work represents a contribution to advance knowledge in the remote detection of algae bloom dynamics and improve monitoring capabilities of inland water bodies.

Assessing the origin of a massive cyanobacterial bloom in the Río de la Plata (2019): Towards an early warning system.

The Río de la Plata estuary drains the second largest river basin of South America. The occurrence of frequent cyanobacterial blooms of the Microcystis and Dolichospermum complex in the Uruguayan coast are associated with high flows of Uruguay River due to rainy years. In summer 2019, a massive cyanobacterial bloom reached up to the Uruguayan Atlantic coast. This study seeks to unveil the origin and the environmental conditions that favored the occurrence of the last cyanobacterial bloom in the Río de la Plata, and to contribute with the development of an early warning system of cyanobacterial scum on Montevideo beaches. A complementary approach was applied with Sentinel-2 imagery, environmental data of monitoring programs of Salto Grande Reservoir and Montevideo beaches, hydro-meteorological information, and hydroelectric dam operation. Images were analyzed with the Normalized Difference Chlorophyll Index (NDCI), which allowed evaluating several water bodies within the same ranges. Positive anomalous rainfall increased river flows, particularly that of Uruguay and Negro rivers, which caused the opening of the dam spillways. NDCI maps showed that areas with high values (NDCI>0.06) in Salto Grande reservoir kept a similar surface area before and after the prolonged overflow period (8.7-7.8 km2, before and after). In the Río Negro reservoirs, however, NDCI>0.06 coverage remarkably changed (62.5 km2, Palmar reservoir), with a subsequent 56-fold reduction in the post-discharge of surface water. Twenty days after opening the spillways, Montevideo beaches were closed to swimming and the NDCI>0.06 surface reached 51.7 km2 in the Río de la Plata coast. The dynamics of NDCI areas, the downstream bloom discharge, and the predicted Río de la Plata residual currents, suggest that the cyanobacterial bloom originated in the Negro River (Palmar reservoir). This bloom event was one of the worst that occurred in the Río de la Plata in last 20 years, circulated along the Uruguayan sub-corridor to the Atlantic coast along 690 km from its origin, and lasted three months on Montevideo coast. This is the first study that addresses the impact of cyanobacterial blooms from the Negro River reservoirs on the Río de la Plata estuary. Therefore, the Negro River basin is where the main efforts should be directed to mitigate massive cyanobacterial blooms.

Biogeography of the cyanobacterium Raphidiopsis (Cylindrospermopsis) raciborskii: Integrating genomics, phylogenetic and toxicity data.

Raphidiopsis (Cylindrospermopsis) raciborskii, a globally distributed bloom-forming cyanobacterium, produces either the cytotoxin cylindrospermopsin (CYL) in Oceania, Asia and Europe or the neurotoxin saxitoxin (STX) and analogues (paralytic shellfish poison, PSP) in South America (encoded by sxt genetic cluster) and none of them in Africa. Nevertheless, this particular geographic pattern is usually overlooked in current hypotheses about the species dispersal routes. Here, we combined genomics, phylogenetic analyses, toxicity data and a literature survey to unveil the evolutionary history and spread of the species. Phylogenies based on 354 orthologous genes from all the available genomes and ribosomal ITS sequences of the taxon showed two well-defined clades: the American, having the PSP producers; and the Oceania/Europe/Asia, including the CYL producers. We propose central Africa as the original dispersion center (non-toxic populations), reaching North Africa and North America (in former Laurasia continent). The ability to produce CYL probably took place in populations that advanced to sub-Saharan Africa and then to Oceania and South America. According to the genomic context of the sxt cluster found in PSP-producer strains, this trait was acquired once by horizontal transfer in South America, where the ability to produce CYL was lost.

Nitrogen availability facilitates phosphorus acquisition by bloom-forming cyanobacteria.

Cyanobacterial blooms are threatening freshwater ecosystems. The physiological basis involved in the onset of cyanobacterial bloom is fundamental to advance in bloom predictions. Generally, cyanobacteria grow until the availability of nitrogen (N), phosphorus (P) or both nutrients becomes limited. Population survival may depend on physiological adjustments to nutrient deficiency as well as on the efficient use of episodic N and P inputs. This study investigated the effect of N inputs on phosphate uptake affinity and activity of N-deficient bloom-forming cyanobacteria. Lake samples dominated by filamentous cyanobacteria were preincubated with and without nitrate addition, and the uptake of [32P] phosphate pulses was measured in the following days. Phosphate uptake kinetics were analyzed with a flow-force model that provides the threshold concentration, reflecting phosphate uptake affinity, and the membrane conductivity coefficient that corresponds to the activity of uptake systems. After 24 h of nitrate preincubation, phosphate uptake kinetics showed a progressive increase in affinity (nanomolar [Pe]A) and activity (25-fold) concomitant with cyanobacterial growth. It was demonstrated that the alleviation of N-deficiency by N inputs boosts the activation of phosphate uptake systems of non-N2-fixing cyanobacteria to sustain growth. Therefore, reduction of dissolved inorganic N levels in lakes is also mandatory to limit cyanobacterial phosphate uptake affinity and activity capabilities.

Influence of nitrogen availability on the expression of genes involved in the biosynthesis of saxitoxin and analogs in Cylindrospermopsis raciborskii.

The development of cyanobacterial blooms in inland aquatic ecosystems is greatly promoted by nutrient availability, especially nitrogen and phosphorous. When blooms are dominated by toxigenic species the harmful effects of nutrient loading becomes particularly relevant. Among toxic species, Cylindrospermopsis raciborskii found in South American ecosystems is characterized by the production of saxitoxin and analogs (Paralytic Shellfish Poisoning, PSP), for which the factors that trigger their production have not been elucidated. In this study, the effect of nitrate availability on the relative transcript abundance of two genes (sxtU and sxtI), both involved in different steps of PSP biosynthetic pathway, was addressed in C. raciborskii MVCC19 by qPCR. The relative transcript abundance of both genes significantly increased from the beginning to the end of growth, independent of nitrate availability in the culture medium. Differences between the genes in terms of the levels of relative expression were also found, implying that during growth in nitrate-rich or nitrate-deprived conditions C. raciborskii MVCC19 has the ability to produce different kind of PSP molecules. The presence of nifH transcripts in the nitrogen-depleted treatment confirmed that in the absence of nitrate C. raciborskii fixed atmospheric N2. Moreover, after transferring filaments to nitrate-rich conditions the synthesis of nifH mRNA continued for few hours, suggesting that cell adjustments enabling the utilization of soluble nitrogen sources are not immediate. Our results show that biosynthesis of saxitoxin and analogs in C. raciborskii is not related to nitrate availability, but rather is linked to cyanobacteria growth rate.

What drives the distribution of the bloom-forming cyanobacteria Planktothrix agardhii and Cylindrospermopsis raciborskii?

The cyanobacteria Planktothrix agardhii and Cylindrospermopsis raciborskii are bloom-forming species common in eutrophic freshwaters. These filamentous species share certain physiological traits which imply that they might flourish under similar environmental conditions. We compared the distribution of the two species in a large database (940 samples) covering different climatic regions and the Northern and Southern hemispheres, and carried out laboratory experiments to compare their morphological and physiological responses. The environmental ranges of the two species overlapped with respect to temperature, light and total phosphorus (TP); however, they responded differently to environmental gradients; C. raciborskii biovolume changed gradually while P. agardhii shifted sharply from being highly dominated to a rare component of the phytoplankton. As expected, P. agardhii dominates the phytoplankton with high TP and low light availability conditions. Contrary to predictions, C. raciborskii succeeded in all climates and at temperatures as low as 11 °C. Cylindrospermopsis raciborskii had higher phenotypic plasticity than P. agardhii in terms of pigments, individual size and growth rates. We conclude that the phenotypic plasticity of C. raciborskii could explain its ongoing expansion to temperate latitudes and suggest its future predominance under predicted climate-change scenarios.

Adaptive phosphate uptake behaviour of phytoplankton to environmental phosphate fluctuations.

When phytoplankton growth in lakes is limited by the available phosphate, the external phosphate concentration fluctuates around a threshold value at which available energy is insufficient to drive phosphate incorporation into a polyphosphate pool. As a result, occasional increases in the external concentration are experienced by phytoplankton as a series of phosphate pulses. Based on [(32) P] phosphate uptake experiments with lake phytoplankton, we show that a community is able to process information about the experienced pattern of phosphate pulses via a complex regulation of the kinetic and energetic properties of cellular phosphate uptake systems. As a result, physiological adaptation to alterations of ambient phosphate concentration depends on the pattern of phosphate fluctuations to which the community had been exposed during its previous growth. In this process, the entire community exhibits coherent uptake behaviour with respect to a common threshold value. Thereby, different threshold values result from different antecedent pulse patterns, apparently unrestrained by the amount of previously stored phosphate. The coherent behaviour observed contradicts the basic assumptions of the competitive exclusion principle and provides an alternative perspective for explaining the paradoxical coexistence of many phytoplankton species.