Catherine Legrand

Allelopathy in phytoplankton - biochemical, ecological and evolutionary aspects

Abstract It is considered self-evident that chemical interactions are a component of competition in terrestrial systems, but they are largely unknown in aquatic systems. In this review, we propose that chemical interactions, specifically allelopathy, are an important part of phytoplankton competition. Allelopathy, as defined here, applies only to the inhibitory effects of secondary metabolites produced by one species on the growth or physiological function of another phytoplankton species. A number of approaches are used to study allelopathy, but there is no standard methodology available. One of the methods used is cross-culturing, in which the cell-free filtrate of a donor alga is added to the medium of the target species. Another is to study the effect of cell extracts of unknown constituents, isolated exudates or purified allelochemicals on the growth of other algal species. There is a clear lack of controlled field experiments because few allelochemicals have been identified. Molecular methods will be important in future to study the expression and regulation of allelochemicals. Most of the identified allelochemicals have been described for cyanobacteria but some known toxins marine dinoflagellates and freshwater cyanobacteria also have an allelochemical effect. The mode of action of allelochemicals spans a wide range. The most common effect is to cause cell lysis, blistering, or growth inhibition. The factors that affect allelochemical production have not been studied much, although nutrient limitation, pH, and temperature appear to have an effect. The evolutionary aspects of allelopathy remain largely unknown, but we hypothesize that the producers of allelochemicals should gain a competitive advantage over other phytoplankton. Finally, we discuss the possibility of using allelochemicals to combat harmful algal blooms (HABs). Allelopathic agents are used for biological control in agriculture, e.g. green manures to control soil diseases in Australia, but they have not yet been applied in the context of HABs. We suggest that phytoplankton allelochemicals have the potential for management of HABs in localized areas.

The Relevance of Marine Chemical Ecology to Plankton and Ecosystem Function: An Emerging Field

Marine chemical ecology comprises the study of the production and interaction of bioactive molecules affecting organism behavior and function. Here we focus on bioactive compounds and interactions associated with phytoplankton, particularly bloom-forming diatoms, prymnesiophytes and dinoflagellates. Planktonic bioactive metabolites are structurally and functionally diverse and some may have multiple simultaneous functions including roles in chemical defense (antipredator, allelopathic and antibacterial compounds), and/or cell-to-cell signaling (e.g., polyunsaturated aldehydes (PUAs) of diatoms). Among inducible chemical defenses in response to grazing, there is high species-specific variability in the effects on grazers, ranging from severe physical incapacitation and/or death to no apparent physiological response, depending on predator susceptibility and detoxification capability. Most bioactive compounds are present in very low concentrations, in both the producing organism and the surrounding aqueous medium. Furthermore, bioactivity may be subject to synergistic interactions with other natural and anthropogenic environmental toxicants. Most, if not all phycotoxins are classic secondary metabolites, but many other bioactive metabolites are simple molecules derived from primary metabolism (e.g., PUAs in diatoms, dimethylsulfoniopropionate (DMSP) in prymnesiophytes). Producing cells do not seem to suffer physiological impact due to their synthesis. Functional genome sequence data and gene expression analysis will provide insights into regulatory and metabolic pathways in producer organisms, as well as identification of mechanisms of action in target organisms. Understanding chemical ecological responses to environmental triggers and chemically-mediated species interactions will help define crucial chemical and molecular processes that help maintain biodiversity and ecosystem functionality.

Local Environmental Conditions Shape Generalist But Not Specialist Components of Microbial Metacommunities in the Baltic Sea

Marine microbes exhibit biogeographical patterns linked with fluxes of matter and energy. Yet, knowledge of the mechanisms shaping bacterioplankton community assembly across temporal scales remains poor. We examined bacterioplankton 16S rRNA gene fragments obtained from Baltic Sea transects to determine phylogenetic relatedness and assembly processes coupled with niche breadth. Communities were phylogenetically more related over time than expected by chance, albeit with considerable temporal variation. Hence, habitat filtering, i.e., local environmental conditions, rather than competition structured bacterioplankton communities in summer but not in spring or autumn. Species sorting (SS) was the dominant assembly process, but temporal and taxonomical variation in mechanisms was observed. For May communities, Cyanobacteria, Actinobacteria, Alpha- and Betaproteobacteria exhibited SS while Bacteroidetes and Verrucomicrobia were assembled by SS and mass effect. Concomitantly, Gammaproteobacteria were assembled by the neutral model and patch dynamics. Temporal variation in habitat filtering and dispersal highlights the impact of seasonally driven reorganization of microbial communities. Typically abundant Baltic Sea populations such as the NS3a marine group (Bacteroidetes) and the SAR86 and SAR11 clade had the highest niche breadth. The verrucomicrobial Spartobacteria population also exhibited high niche breadth. Surprisingly, variation in bacterioplankton community composition was regulated by environmental factors for generalist taxa but not specialists. Our results suggest that generalists such as NS3a, SAR86, and SAR11 are reorganized to a greater extent by changes in the environment compared to specialists and contribute more strongly to determining overall biogeographical patterns of marine bacterial communities.

Disentangling seasonal bacterioplankton population dynamics by high‐frequency sampling

Multiyear comparisons of bacterioplankton succession reveal that environmental conditions drive community shifts with repeatable patterns between years. However, corresponding insight into bacterioplankton dynamics at a temporal resolution relevant for detailed examination of variation and characteristics of specific populations within years is essentially lacking. During 1 year, we collected 46 samples in the Baltic Sea for assessing bacterial community composition by 16S rRNA gene pyrosequencing (nearly twice weekly during productive season). Beta-diversity analysis showed distinct clustering of samples, attributable to seemingly synchronous temporal transitions among populations (populations defined by 97% 16S rRNA gene sequence identity). A wide spectrum of bacterioplankton dynamics was evident, where divergent temporal patterns resulted both from pronounced differences in relative abundance and presence/absence of populations. Rates of change in relative abundance calculated for individual populations ranged from 0.23 to 1.79 day(-1) . Populations that were persistently dominant, transiently abundant or generally rare were found in several major bacterial groups, implying evolution has favoured a similar variety of life strategies within these groups. These findings suggest that high temporal resolution sampling allows constraining the timescales and frequencies at which distinct populations transition between being abundant or rare, thus potentially providing clues about physical, chemical or biological forcing on bacterioplankton community structure.

Ocean urea fertilization for carbon credits poses high ecological risks

The proposed plan for enrichment of the Sulu Sea, Philippines, a region of rich marine biodiversity, with thousands of tonnes of urea in order to stimulate algal blooms and sequester carbon is flawed for multiple reasons. Urea is preferentially used as a nitrogen source by some cyanobacteria and dinoflagellates, many of which are neutrally or positively buoyant. Biological pumps to the deep sea are classically leaky, and the inefficient burial of new biomass makes the estimation of a net loss of carbon from the atmosphere questionable at best. The potential for growth of toxic dinoflagellates is also high, as many grow well on urea and some even increase their toxicity when grown on urea. Many toxic dinoflagellates form cysts which can settle to the sediment and germinate in subsequent years, forming new blooms even without further fertilization. If large-scale blooms do occur, it is likely that they will contribute to hypoxia in the bottom waters upon decomposition. Lastly, urea production requires fossil fuel usage, further limiting the potential for net carbon sequestration. The environmental and economic impacts are potentially great and need to be rigorously assessed.

Metagenome-assembled genomes uncover a global brackish microbiome

BackgroundMicrobes are main drivers of biogeochemical cycles in oceans and lakes. Although the genome is a foundation for understanding the metabolism, ecology and evolution of an organism, few bacterioplankton genomes have been sequenced, partly due to difficulties in cultivating them.ResultsWe use automatic binning to reconstruct a large number of bacterioplankton genomes from a metagenomic time-series from the Baltic Sea, one of world’s largest brackish water bodies. These genomes represent novel species within typical freshwater and marine clades, including clades not previously sequenced. The genomes’ seasonal dynamics follow phylogenetic patterns, but with fine-grained lineage-specific variations, reflected in gene-content. Signs of streamlining are evident in most genomes, and estimated genome sizes correlate with abundance variation across filter size fractions. Comparing the genomes with globally distributed metagenomes reveals significant fragment recruitment at high sequence identity from brackish waters in North America, but little from lakes or oceans. This suggests the existence of a global brackish metacommunity whose populations diverged from freshwater and marine relatives over 100,000 years ago, long before the Baltic Sea was formed (8000 years ago). This markedly contrasts to most Baltic Sea multicellular organisms, which are locally adapted populations of freshwater or marine counterparts.ConclusionsWe describe the gene content, temporal dynamics and biogeography of a large set of new bacterioplankton genomes assembled from metagenomes. We propose that brackish environments exert such strong selection that lineages adapted to them flourish globally with limited influence from surrounding aquatic communities.

Transplant experiments uncover Baltic Sea basin-specific responses in bacterioplankton community composition and metabolic activities

Anthropogenically induced changes in precipitation are projected to generate increased river runoff to semi-enclosed seas, increasing loads of terrestrial dissolved organic matter and decreasing salinity. To determine how bacterial community structure and functioning adjust to such changes, we designed microcosm transplant experiments with Baltic Proper (salinity 7.2) and Bothnian Sea (salinity 3.6) water. Baltic Proper bacteria generally reached higher abundances than Bothnian Sea bacteria in both Baltic Proper and Bothnian Sea water, indicating higher adaptability. Moreover, Baltic Proper bacteria growing in Bothnian Sea water consistently showed highest bacterial production and beta-glucosidase activity. These metabolic responses were accompanied by basin-specific changes in bacterial community structure. For example, Baltic Proper Pseudomonas and Limnobacter populations increased markedly in relative abundance in Bothnian Sea water, indicating a replacement effect. In contrast, Roseobacter and Rheinheimera populations were stable or increased in abundance when challenged by either of the waters, indicating an adjustment effect. Transplants to Bothnian Sea water triggered the initial emergence of particular Burkholderiaceae populations, and transplants to Baltic Proper water triggered Alteromonadaceae populations. Notably, in the subsequent re-transplant experiment, a priming effect resulted in further increases to dominance of these populations. Correlated changes in community composition and metabolic activity were observed only in the transplant experiment and only at relatively high phylogenetic resolution. This suggested an importance of successional progression for interpreting relationships between bacterial community composition and functioning. We infer that priming effects on bacterial community structure by natural episodic events or climate change induced forcing could translate into long-term changes in bacterial ecosystem process rates.

Unscrambling Cyanobacteria Community Dynamics Related to Environmental Factors

Future climate scenarios in the Baltic Sea project an increase of cyanobacterial bloom frequency and duration, attributed to eutrophication and climate change. Some cyanobacteria can be toxic and their impact on ecosystem services is relevant for a sustainable sea. Yet, there is limited understanding of the mechanisms regulating cyanobacterial diversity and biogeography. Here we unravel successional patterns and changes in cyanobacterial community structure using a 2-year monthly time- series during the productive season in a 100 km coastal-offshore transect using microscopy and high-throughput sequencing of 16S rRNA gene fragments. A total of 565 cyanobacterial OTUs were found, of which 231 where filamentous/colonial and 334 picocyanobacterial. Spatial differences in community structure between coastal and offshore waters were minor. An “epidemic population structure” (dominance of asingle cluster) was found for Aphanizomenon/Dolichospermum within the filamentous/colonial cyanobacterial community. In summer, this clusters imultaneously occurred with opportunistic clusters/OTUs, e.g., Nodularia spumigena and Pseudanabaena. Picocyanobacteria, Synechococcus/Cyanobium, formeda consistent but highly diverse group. Overall, the potential drivers structuring summer cyanobacterial communities were temperature and salinity. However, the different responses to environmental factors among and within genera suggest high niche specificity for individual OTUs. The recruitment and occurrence of potentially toxic filamentous/colonial clusters was likely related to disturbance such as mixing events and short-term shifts in salinity, and not solely dependent on increasing temperature and nitrogen-limiting conditions. Nutrients did not explain further the changes in cyanobacterial community composition. Novel occurrence patterns were identified as a strong seasonal succession revealing a tight coupling between the emergence of opportunistic picocynobacteria and the bloom offilamentous/colonialclusters. These findings highlight that if environmental conditions can partially explain the presence of opportunistic picocyanobacteria, microbial and trophic interactions with filamentous/colonial cyanobacteria should also be considered as potential shaping factors for single-celled communities. Regional climate change scenarios in the Baltic Sea predict environmental shifts leading to higher temperature and lower salinity; conditions identified here as favorable for opportunistic filamentous/colonial cyanobacteria. Altogether, the diversity and complexity of cyanobacterial communities reported here is far greater than previously known, emphasizing the importance of microbial interactions between filamentous and picocyanobacteria in the context of environmental disturbances.

Spatio-Temporal Interdependence of Bacteria and Phytoplankton during a Baltic Sea Spring Bloom

In temperate systems, phytoplankton spring blooms deplete inorganic nutrients and are major sources of organic matter for the microbial loop. In response to phytoplankton exudates and environmental factors, heterotrophic microbial communities are highly dynamic and change their abundance and composition both on spatial and temporal scales. Yet, most of our understanding about these processes comes from laboratory model organism studies, mesocosm experiments or single temporal transects. Spatial-temporal studies examining interactions of phytoplankton blooms and bacterioplankton community composition and function, though being highly informative, are scarce. In this study, pelagic microbial community dynamics (bacteria and phytoplankton) and environmental variables were monitored during a spring bloom across the Baltic Proper (two cruises between North Germany to Gulf of Finland). To test to what extent bacterioplankton community composition relates to the spring bloom, we used next generation amplicon sequencing of the 16S rRNA gene, phytoplankton diversity analysis based on microscopy counts and population genotyping of the dominating diatom Skeletonema marinoi. Several phytoplankton bloom related and environmental variables were identified to influence bacterial community composition. Members of Bacteroidetes and Alphaproteobacteria dominated the bacterial community composition but the bacterial groups showed no apparent correlation with direct bloom related variables. The less abundant bacterial phyla Actinobacteria, Planctomycetes, and Verrucomicrobia, on the other hand, were strongly associated with phytoplankton biomass, diatom:dinoflagellate ratio, and colored dissolved organic matter (cDOM). Many bacterial operational taxonomic units (OTUs) showed high niche specificities. For example, particular Bacteroidetes OTUs were associated with two distinct genetic clusters of S. marinoi. Our study revealed the complexity of interactions of bacterial taxa with inter- and intraspecific genetic variation in phytoplankton. Overall, our findings imply that biotic and abiotic factors during spring bloom influence bacterial community dynamics in a hierarchical manner.

Rapid clean-up and effective sample preparation procedure for unambiguous determination of the cyclic peptides microcystin and nodularin

SummaryA new sample preparation strategy has been established to improve the identification and determination of nodularin and microcystins. The sample preparation consisted of enrichment of the analytes by solid phase extraction with C18 cartridges followed by clean-up of the enriched raw extracts by high performance size exclusion gel permeation chromatography. In contrast to established clean-up procedures based on polarity, related distribution of microcystins and nodularin in non-miscible phases (e. g. a C18 cartridge as stationary phase and a water-containing eluent as mobile phase) this strategy separates microcystins from interfering compounds by molecular size differences.The sample preparation procedure can be automated easily and was validated for both water samples as well as raw extracts of algal cells. The method was success-fully applied during an experiment with natural algae communities from the Baltic Sea to investigate the influence of different nutrient limitations on toxicity ofNodularia sp...