How do seasonal temperature variations influence interplay between toxic and non-toxic cyanobacterial blooms? Evidence from modeling and experimental data.

Summer cyanobacterial blooms exhibit a dynamic interplay between toxic and non-toxic genotypes, significantly influencing the cyanotoxin levels within a lake. The challenge lies in accurately predicting these toxin concentrations due to the significant temporal fluctuations in the proportions of toxic and non-toxic genotypes. Typically, the toxic genotypes dominate during the early and late summer periods, while the non-toxic variants prevail in mid-summer. To dissect this phenomenon, we propose a model that accounts for the competitive interaction between toxic and non-toxic genotypes, as well as seasonal temperature variations. Our numerical simulations suggest that the optimal temperature of the toxic genotypes is lower than that of the optimal temperatures of the non-toxic counterparts. This difference of optimal temperature may potentially contribute to explain the dominance of toxic genotypes at the early and late summer periods, situation often observed in the field. Experimental data from the laboratory align qualitatively with our simulation results, enabling a better understanding of complex interplays between toxic and non-toxic cyanobacteria.

Cyanosphere Dynamic During Dolichospermum Bloom: Potential Roles in Cyanobacterial Proliferation.

Under the effect of global change, management of cyanobacterial proliferation becomes increasingly pressing. Given the importance of interactions within microbial communities in aquatic ecosystems, a handful of studies explored the potential relations between cyanobacteria and their associated bacterial community (i.e., cyanosphere). Yet, most of them specifically focused on the ubiquitous cyanobacteria Microcystis, overlooking other genera. Here, based on 16s rDNA metabarcoding analysis, we confirmed the presence of cyanosphere representing up to 30% of the total bacterial community diversity, during bloom episode of another preponderant cyanobacterial genus, Dolichospermum. Moreover, we highlighted a temporal dynamic of this cyanosphere. A sPLS-DA model permits to discriminate three important dates and 220 OTUs. With their affiliations, we were able to show how these variations potentially imply a turnover in ecological functions depending on bloom phases. Although more studies are necessary to quantify the impacts of these variations, we argue that cyanosphere can have an important, yet underestimated, role in the modulation of cyanobacterial blooms.

ConnecSenS, a Versatile IoT Platform for Environment Monitoring: Bring Water to Cloud.

Climate change is having an increasingly rapid impact on ecosystems and particularly on the issue of water resources. The Internet of Things and communication technologies have now reached a level of maturity that allows sensors to be deployed more easily on sites to monitor them. The communicating node based on LoRaWAN technology presented in this article is open and allows the interfacing of numerous sensors for designing long-term environmental monitoring systems of isolated sites. The data integration in the cloud is ensured by a workflow driving the storage and indexing of data, allowing a simple and efficient use of the data for different users (scientists, administration, citizens) through specific dashboards and extractions. This article presents this infrastructure through environmental monitoring use cases related to water resources.

Higher sensitivity to hydrogen peroxide and light stress conditions of the microcystin producer Microcystis aeruginosa sp PCC7806 compared to non-producer strains.

The increasing incidence of cyanobacterial blooms with their associated production of cyanotoxins lead managers of aquatics systems to control their biomass to limit the health risk. Among the variety of existing treatment approaches, hydrogen peroxide (H2O2) shows increasing use but the effects of environmental parameters on its effectiveness are still not completely known. With the aim to assess the efficiency of H2O2 treatments in the control of cyanobacterial blooms and decrease toxic risk, we tested three Microcystis strains according to their ability to produce cyanotoxins (a microcystin-producing, non-microcystin-producing and mcyB-knockout mutant). Photochemical efficiency, percentage of living cells and microcystin cell content were compared under various hydrogen peroxide concentrations coupled with stress conditions encountered during the life cycle of cyanobacteria as darkness and high light. The microcystin-producing strain appeared the more sensitive to hydrogen peroxide treatment and to light condition, probably due to a lower rate of repair of Photo System II (PSII). We also highlighted various responses of PSII activity according to Microcystis strains which could partly explain the shift of dominant genotypes often occurring during a bloom event. Our results confirm the link between light and microcystin content and variations of microcystin contents appear as a consequence of photosynthetic activity. These findings could be of particular interest regarding water quality management, especially the use of H2O2 as a potential algaecide which seems to be more effective to use during periods of high light.

Global co-occurrence of methanogenic archaea and methanotrophic bacteria in Microcystis aggregates.

Global warming and eutrophication contribute to the worldwide increase in cyanobacterial blooms, and the level of cyanobacterial biomass is strongly associated with rises in methane emissions from surface lake waters. Hence, methane-metabolizing microorganisms may be important for modulating carbon flow in cyanobacterial blooms. Here, we surveyed methanogenic and methanotrophic communities associated with floating Microcystis aggregates in 10 lakes spanning four continents, through sequencing of 16S rRNA and functional marker genes. Methanogenic archaea (mainly Methanoregula and Methanosaeta) were detectable in 5 of the 10 lakes and constituted the majority (~50%-90%) of the archaeal community in these lakes. Three of the 10 lakes contained relatively more abundant methanotrophs than the other seven lakes, with the methanotrophic genera Methyloparacoccus, Crenothrix, and an uncultured species related to Methylobacter dominating and nearly exclusively found in each of those three lakes. These three are among the five lakes in which methanogens were observed. Operational taxonomic unit (OTU) richness and abundance of methanotrophs were strongly positively correlated with those of methanogens, suggesting that their activities may be coupled. These Microcystis-aggregate-associated methanotrophs may be responsible for a hitherto overlooked sink for methane in surface freshwaters, and their co-occurrence with methanogens sheds light on the methane cycle in cyanobacterial aggregates.

The global Microcystis interactome.

Bacteria play key roles in the function and diversity of aquatic systems, but aside from study of specific bloom systems, little is known about the diversity or biogeography of bacteria associated with harmful cyanobacterial blooms (cyanoHABs). CyanoHAB species are known to shape bacterial community composition and to rely on functions provided by the associated bacteria, leading to the hypothesized cyanoHAB interactome, a coevolved community of synergistic and interacting bacteria species, each necessary for the success of the others. Here, we surveyed the microbiome associated with Microcystis aeruginosa during blooms in 12 lakes spanning four continents as an initial test of the hypothesized Microcystis interactome. We predicted that microbiome composition and functional potential would be similar across blooms globally. Our results, as revealed by 16S rRNA sequence similarity, indicate that M. aeruginosa is cosmopolitan in lakes across a 280° longitudinal and 90° latitudinal gradient. The microbiome communities were represented by a wide range of operational taxonomic units and relative abundances. Highly abundant taxa were more related and shared across most sites and did not vary with geographic distance, thus, like Microcystis, revealing no evidence for dispersal limitation. High phylogenetic relatedness, both within and across lakes, indicates that microbiome bacteria with similar functional potential were associated with all blooms. While Microcystis and the microbiome bacteria shared many genes, whole-community metagenomic analysis revealed a suite of biochemical pathways that could be considered complementary. Our results demonstrate a high degree of similarity across global Microcystis blooms, thereby providing initial support for the hypothesized Microcystis interactome.

Akinetes and ancient DNA reveal toxic cyanobacterial recurrences and their potential for resurrection in a 6700-year-old core from a eutrophic lake.

In order to evaluate the recurrence of toxic cyanobacterial blooms and to determine the survival capabilities of the resistance cells through time, a sedimentary core spanning 6700 years was drilled in the eutrophic Lake Aydat. A multiproxy approach (density, magnetic susceptibility, XRF, pollen and non-pollen palynomorph analyses), was used initially to determine the sedimentation model and the land uses around the lake. Comparison with the akinete count revealed that Nostocales cyanobacteria have been present in Lake Aydat over a six thousand year period. This long-term cyanobacterial recurrence also highlights the past presence of both the anaC and mcyB genes, involved in anatoxin-a and microcystin biosynthesis, respectively, throughout the core. The first appearance of cyanobacteria seems to be linked to the natural damming of the river, while the large increase in akinete density around 1800 cal.yr BP can be correlated with the intensification of human activities (woodland clearance, crop planting, grazing, etc.) in the catchment area of the lake, and marks the beginning of a long period of eutrophication. This first investigation into the viability and germination potential of cyanobacteria over thousands of years reveals the ability of intact akinetes to undergo cell divisions even after 1800 years of sedimentation, which is 10 times longer than previously observed. This exceptional cellular resistance, coupled with the long-term eutrophic conditions of this lake, could partly explain the past and current recurrences of cyanobacterial proliferations.

Akinetes May Be Representative of Past Nostocalean Blooms: a Case Study of Their Benthic Spatiotemporal Distribution and Potential for Germination in a Eutrophic Lake.

Monitoring of water and surface sediment in a French eutrophic lake (Lake Aydat) was carried out over a 2-year period in order to determine whether akinetes in sediment could be representative of the most recent bloom and to estimate their germination potential. Sediment analysis revealed two akinete species, Dolichospermum macrosporum and Dolichospermumflos-aquae, present in the same proportions as observed for the pelagic populations. Moreover, similar spatial patterns observed for vegetative cells in the water column and akinete distributions in the sediment suggest that akinetes in the sediment may be representative of the previous bloom. However, the relationship between akinetes in the sediment and vegetative cells in the water column was not linear, and other factors may interfere. For example, our results highlighted horizontal transport of akinetes during the winter. The benthic overwinter phase did not seem to influence the percentages of intact akinetes, which remained stable at approximately 7% and 60% for D. macrosporum and D. flos-aquae, respectively. These percentages may thus be the result of processes that occurred in the water column. The intact overwintering akinetes showed germination rates of up to 90% after 72 h for D. flos-aquae or 144 h for D. macrosporum The difference in akinete germination rates between these two species demonstrates different ecological strategies, which serve to expand the window for germination in time and space and thus optimize colonization of the water column by nostocalean cyanobacteria.IMPORTANCE Cyanobacteria have the ability to proliferate and to form blooms. These blooms can then affect the local ecology, health, and economy. The akinete, a resistant cell type that persists in sediment, is an important intermediate phase between previous and future blooms. We monitored the water column and the surface sediment of a French eutrophic lake (Lake Aydat) to investigate the relationship between vegetative cells in the water column and akinetes in the sediment. This study focused on the characterization of spatiotemporal akinete distributions, cellular integrity, and germination potential. Species-specific ecological strategies were highlighted and may partly explain the temporal succession of species in the water column. Akinetes may also be used to understand past nostocalean blooms and to predict future ones.

Benthic Archives Reveal Recurrence and Dominance of Toxigenic Cyanobacteria in a Eutrophic Lake over the Last 220 Years.

Akinetes are resistant cells which have the ability to persist in sediment for several decades. We have investigated the temporal distribution of akinetes of two species, Dolichospermum macrosporum and Dolichospermum flos-aquae, in a sediment core sampled in Lake Aydat (France), which covers 220 years. The upper part, from 1907 to 2016, the number of akinetes fluctuated but stayed at high concentrations, especially for D. macrosporum in surface sediment (with the maximal value close to 6.105 akinetes g DW-1 of sediment), suggesting a recurrence of blooms of this species which was probably closely related to anthropic eutrophication since the 1960s. Before 1907, the abundance of akinetes of both species was very low, suggesting only a modest presence of these cyanobacteria. In addition, the percentage of intact akinetes was different for each species, suggesting different ecological processes in the water column. This percentage also decreased with depth, revealing a reduction in germination potential over time. In addition, biosynthetic genes of anatoxin-a (anaC) and microcystin (mcyA) were detected. First results show a high occurrence of mcyA all down the core. In contrast, anaC gene was mostly detected in the surface sediment (since the 1980s), revealing a potentially more recent occurrence of this cyanotoxin in Lake Aydat which may be associated with the recurrence of blooms of D. macrosporum and thus with anthropic activities.

Spatial and temporal changes of parasitic chytrids of cyanobacteria.

Parasitism is certainly one of the most important driving biotic factors of cyanobacterial blooms which remains largely understudied. Among these parasites, fungi from the phylum Chytridiomycota (i.e. chytrids) are the only eukaryotic microorganisms infecting cyanobacteria. Here, we address spatiotemporal dynamics of the cyanobacterial host Dolichospermum macrosporum (syn. Anabaena macrospora) and its associated chytrid parasites, Rhizosiphon spp., in an eutrophic lake by studying spatial (vertical, horizontal) and temporal (annual and inter-annual) variations. Our results show homogenous chytrid infection patterns along the water column and across sampling stations. However, the prevalence of infection presented drastic changes with time, at both intra- and inter-annual scales. In 2007, a maximum of 98% of vegetative cells were infected by R. crassum whereas this fungal species was not reported seven years later. In opposite, R. akinetum, a chytrid infecting only akinetes, increased its prevalence by 42% during the same period. High chytrid infection rate on the akinetes might have sizeable consequences on host recruitment (and proliferation) success from year to year, as supported by the recorded inter-annual host dynamics (affecting also the success of other chytrid parasites). The spatial homogenous chytrid infection on this cyanobacterium, coupled to both seasonal and inter-annual changes indicates that time, rather than space, controls such highly dynamic host-parasite relationships.

Molecular tools to detect anatoxin-a genes in aquatic ecosystems: Toward a new nested PCR-based method.

Over the last few decades, cyanobacterial mass occurrence has become a recurrent feature of aquatic ecosystems. This has led to ecosystem exposure and health hazards associated with cyanotoxin production. The neurotoxin anatoxin-a and its homologs can be synthesized by benthic cyanobacterial species in lotic systems, but also by planktonic lacustrine species such as Dolichospermum (also known as Anabaena). However, only a few studies have focused on anatoxin-a occurrence and its biosynthesis genes in freshwater lakes. The initial aim of this study was to evaluate the molecular tools available in the literature to detect anatoxin-a biosynthesis genes in lacustrine environments. Having tested different sets of PCR primers, we found that that some sets of primers, such as anxC, were too specific and did not amplify anatoxin-a biosynthesis genes in all producing strains. On the other hand, some sets of primers, such as atxoa, seemed not to be specific enough, amplifying numerous non-specific bands in environmental samples, especially those from sediments. Furthermore, anaC and anaF amplification exhibited different band intensities during electrophoresis, suggesting a high variation in number of gene copies between samples. As a result, we proposed a new nested PCR-based method which considerably improved the amplification of the anaC gene in our environmental samples, eliminating non-specific bands and weak detections. Using this tool, our study also highlighted that anatoxin-a genes are widely distributed throughout freshwater lakes. This suggests the need for further ecological investigations into anatoxin-a in these ecosystems.

Characterization of akinetes from cyanobacterial strains and lake sediment: A study of their resistance and toxic potential.

Nostocalean cyanobacteria are known to proliferate abundantly in eutrophic aquatic ecosystems, and to produce several cyanotoxins, including anatoxin-a. In this study, we investigated both the resistance and toxic potential of the akinetes (resistant cells), using cyanobacterial cultures and akinetes extracted from the sediment of Lake Aydat (France) sampled in the winter and spring. Intact and lysed akinetes were differentiated using a double control based on the autofluorescence of akinetes and SYTOX-green staining. The percentage of resistant akinetes found in several different abiotic stress conditions was highly variable, depending on the species and also on the sampling season. Thus, the resistance of akinetes and their ability to germinate seems to follow a species-specific process, and akinetes can undergo physiologic changes during the sedimentary phase of the Nostocale life cycle. This study also revealed the first evidence of anatoxin-a genes in akinetes, with anaC and anaF genes detected in akinetes from all cyanobacterial producer cultures. The low number of anaC genes, almost exclusively detected using nested PCR, in the sediment at Lake Aydat suggests a limited but existent past population of toxic Nostocales in this lake. Given the key role of akinetes in the annual cycle and subsequent summer proliferation, it can be interesting to integrate the surveillance of akinetes in the management of lakes exposed to recurrent cyanobacterial blooms.

Microbial players involved in the decline of filamentous and colonial cyanobacterial blooms with a focus on fungal parasitism.

In the forthcoming decades, it is widely believed that the dominance of colonial and filamentous bloom-forming cyanobacteria (e.g. Microcystis, Planktothrix, Anabaena and Cylindrospermopsis) will increase in freshwater systems as a combined result of anthropogenic nutrient input into freshwater bodies and climate change. While the physicochemical parameters controlling bloom dynamics are well known, the role of biotic factors remains comparatively poorly studied. Morphology and toxicity often - but not always - limit the availability of cyanobacteria to filter feeding zooplankton (e.g. cladocerans). Filamentous and colonial cyanobacteria are widely regarded as trophic dead-ends mostly inedible for zooplankton, but substantial evidence shows that some grazers (e.g. copepods) can bypass this size constraint by breaking down filaments, making the bloom biomass available to other zooplankton species. A wide range of algicidal bacteria (mostly from the Alcaligenes, Flavobacterium/Cytophaga group and Pseudomonas) and viruses (Podoviridae, Siphoviridae and Myoviridae) may also contribute to bloom control, via their lytic activity underpinned by a diverse array of mechanisms. Fungal parasitism by the Chytridiomycota remains the least studied. While each of these biotic factors has traditionally been studied in isolation, emerging research consistently point to complex interwoven interactions between biotic and environmental factors.

Co-occurrence of microcystin and anatoxin-a in the freshwater lake Aydat (France): Analytical and molecular approaches during a three-year survey.

Cyanobacterial mass occurrence is becoming a growing concern worldwide. They notably pose a threat to water users when cyanotoxins are produced. The aim of this study was to evaluate the occurrence and the dynamics of two cyanotoxins: microcystin (MC) and anatoxin-a (ANTX-a), and of two of the genes responsible for their production (respectively mcyA and anaC) during three consecutive bloom periods (2011, 2012 and 2013) in Lake Aydat (Auvergne, France). MC was detected at all sampling dates, but its concentration showed strong inter- and intra-annual variations. MC content did not correlate with cyanobacterial abundance, nor with any genera taken individually, but it significantly correlated with mcyA gene abundance (R2=0.51; p=0.042). MC content and mcyA gene abundance were maximal when cyanobacterial abundance was low, either at the onset of the bloom or during a trough of biomass. The LC-MS/MS analysis showed the presence of ANTX-a in the 2011 samples. To our knowledge, this is the first report of the presence of this neurotoxin in a French lake. The presence of ANTX-a corresponded to the only year for which Anabaena did not dominate the cyanobacterial community alone, and several cyanobacterial genera were present, including notably Aphanizomenon. anaC gene detection by PCR was not coherent with ANTX-a presence, both gene and toxin were never found for a same sample. This implies that molecular tools to study genes responsible for the production of anatoxin-a are still imperfect and the development of new primers is needed. This study also highlights the need for better monitoring practices that would not necessarily focus only on the peak of cyanobacterial abundance and that would take cyanotoxins other than MC into account.

Genetic diversity along the life cycle of the cyanobacterium Microcystis: highlight on the complexity of benthic and planktonic interactions.

Microcystis is a toxic freshwater cyanobacterium with an annual life cycle characterized by the alternation of a planktonic proliferation stage in summer and a benthic resting stage in winter. Given the importance of both stages for the development and the survival of the population, we investigated the genotypic composition of the planktonic and benthic Microcystis subpopulations from the Grangent reservoir (France) during two distinct proliferation periods. Our results showed a succession of different dominant genotypes in the sediment as well as in the water all along the study periods with some common genotypes to both compartments. Analysis of molecular variance and UniFrac analysis confirmed the similarity between some benthic and planktonic samples, thus evidencing exchanges of genotypes between water and sediment. Thanks to these data, recruitment and sedimentation were proven not to be restricted to spring and autumn, contrary to what was previously thought. Finally, genetic diversity was significantly higher in the sediment than in the water (P < 0.01; Student's t-test). Taken together, our results shed light on the hidden contribution of the benthic compartment in maintaining the genetic diversity of Microcystis populations throughout their annual cycle, which could explain their ecological success in aquatic ecosystems.

The importance of small colonies in sustaining Microcystis population exposed to mixing conditions: an exploration through colony size, genotypic composition and toxic potential.

Microcystis is a toxic colony-forming cyanobacterium, which can bloom in a wide range of freshwater ecosystems. Despite the ecological advantage of the colonial form, few studies have paid attention to the size of Microcystis colonies in the field. With the aim of evaluating the impact of a fluctuating physical environment on the colony size, the genotypic composition and the toxic potential of a Microcystis population, we investigated five different colony size classes of a Microcystis bloom in the Grangent reservoir (France). By sequencing the internal transcribed spacer of the ribosomal operon, we evidenced changes in the genetic structure among size classes in response to environmental change. While similar genotypes were seen in every size class in stable conditions, new dominant genotypes appeared in the smallest colonies (< 160 µm) concomitantly with mixing conditions, strongly suggesting the importance of these colonies in response to disturbances. Moreover, these small colonies played a major role in microcystin production during this bloom, since very high microcystin contents (> 1 pg.cell.(-1)) were found in their cells. These findings indicate that the colony size distribution of a Microcystis population in response to disturbance could be an adaptive strategy that may explain its ecological success in freshwater ecosystems.

A double staining method using SYTOX green and calcofluor white for studying fungal parasites of phytoplankton.

We propose a double staining method based on the combination of two fluorochromes, calcofluor white (CFW; specific chitinous fluorochrome) and SYTOX green (nucleic acid stain), coupled to epifluorescence microscopy for counting, identifying, and investigating the fecundity of parasitic fungi of phytoplankton and the putative relationships established between hosts and their chytrid parasites. The method was applied to freshwater samples collected over two successive years during the terminal period of autumnal cyanobacterial blooms in a eutrophic lake. The study focused on the uncultured host-parasite couple Anabaena macrospora (cyanobacterium) and Rhizosiphon akinetum (Chytridiomycota). Our results showed that up to 36.6% of cyanobacterial akinetes could be parasitized by fungi. Simultaneously, we directly investigated the zoosporic content inside the sporangia and found that both the host size and intensity of infection conditioned the final size and hence fecundity of the chytrids. We found that relationships linking host size, final parasite size, and chytrid fecundity were conserved from year to year and seemed to be host-chytrid couple specific. We concluded that our double staining method was a valid procedure for improving our knowledge of uncultured freshwater phytoplankton-chytrid couples and so of the quantitative ecology of chytrids in freshwater ecosystems.

Fungal parasitism: life cycle, dynamics and impact on cyanobacterial blooms.

Many species of phytoplankton are susceptible to parasitism by fungi from the phylum Chytridiomycota (i.e. chytrids). However, few studies have reported the effects of fungal parasites on filamentous cyanobacterial blooms. To investigate the missing components of bloom ecosystems, we examined an entire field bloom of the cyanobacterium Anabaena macrospora for evidence of chytrid infection in a productive freshwater lake, using a high resolution sampling strategy. A. macrospora was infected by two species of the genus Rhizosiphon which have similar life cycles but differed in their infective regimes depending on the cellular niches offered by their host. R. crassum infected both vegetative cells and akinetes while R. akinetum infected only akinetes. A tentative reconstruction of the developmental stages suggested that the life cycle of R. crassum was completed in about 3 days. The infection affected 6% of total cells (and 4% of akinètes), spread over a maximum of 17% of the filaments of cyanobacteria, in which 60% of the cells could be parasitized. Furthermore, chytrids may reduce the length of filaments of Anabaena macrospora significantly by "mechanistic fragmentation" following infection. All these results suggest that chytrid parasitism is one of the driving factors involved in the decline of a cyanobacteria blooms, by direct mortality of parasitized cells and indirectly by the mechanistic fragmentation, which could weaken the resistance of A. macrospora to grazing.

Short- and long-term dynamics of the toxic potential and genotypic structure in benthic populations of Microcystis.

Microcystis colonies are known to overwinter on the surface of the sediment of freshwater ecosystems. However, little is known about the genotypic and toxicological dynamics of Microcystis populations during this benthic life stage. In this study, we report a two-year-long survey of benthic populations of Microcystis, which had spent from a few days to more than six years in the sediment. In order to avoid any interaction with the planktonic proliferations, we chose two deeply buried benthic populations, which could be easily dated. Quantitative PCR on mcyB gene and protein phosphatase inhibition assays were performed to measure their toxic potential, and their genotypic structure was assessed by Capillary Electrophoresis-Single Strand Conformation Polymorphism (CE-SSCP), based on 16S-23S Intergenic Transcribed Spacer (ITS). The microcystin content of the cells seemed to change sharply during the first few months of benthic survival, whereas this content was low and decreased steadily after several years of benthic life. No genetic selection was observed in either the proportion of potentially toxic clones or the ITS sequences for any of the populations considered. From these results, the benthic life stage of Microcystis appears to preserve the structure and the composition of the population over a far larger time scale than classical overwintering period. Finally, some genotypes were common in both of the benthic populations, even though they originated from planktonic blooms that had developed five years apart, suggesting a major overlap of planktonic proliferations in successive years.

INVOLVEMENT OF MICROCYSTINS AND COLONY SIZE IN THE BENTHIC RECRUITMENT OF THE CYANOBACTERIUM MICROCYSTIS (CYANOPHYCEAE)(1).

The benthic recruitment of Microcystis was simulated in vitro in order to characterize the colonies of Microcystis recruited and to study the impact of intracellular and extracellular microcystins (MCs), and the influence of colony size on the recruitment process. We observed recruitment dynamics consisting of a lag phase followed by a peak and then a return to low recruitment rates, mainly controlled by passive resuspension throughout the experiment, and by physiological processes during the recruitment peak. Ninety-seven percent of the Microcystis colonies recruited were <160 µm in maximum length, and their cells contained much greater amounts of MCs (0.26 ± 0.14 pg eq microcystin leucine-arginine variant [MC-LR] · cell(-1) ) than those in benthic colonies (0.021 ± 0.004 pg eq MC-LR · cell(-1) ). The MC content of recruited Microcystis varied significantly over time and was not related to changes in the proportion of potentially toxic genotypes, determined using real-time PCR. On the other hand, the changes in MC content in the potentially toxic Microcystis recruited were closely and negatively correlated with recruitment dynamics; the lowest MC contents corresponded to high recruitment rates, and the highest MC contents corresponded to low recruitment rates. Thus, depending on temperature and light conditions, these variations are thought to result from the selection of various subpopulations from among the smallest and the most toxic of the initial benthic population. Adding purified MC-LR to experimental treatments led to a decreased recruitment of Microcystis and more specifically of mcyB genotypes.