Raffaella Casotti

Aldehyde suppression of copepod recruitment in blooms of a ubiquitous planktonic diatom

The growth cycle in nutrient-rich, aquatic environments starts with a diatom bloom that ends in mass sinking of ungrazed cells and phytodetritus. The low grazing pressure on these blooms has been attributed to the inability of overwintering copepod populations to track them temporally. We tested an alternative explanation: that dominant diatom species impair the reproductive success of their grazers. We compared larval development of a common overwintering copepod fed on a ubiquitous, early-blooming diatom species with its development when fed on a typical post-bloom dinoflagellate. Development was arrested in all larvae in which both mothers and their larvae were fed the diatom diet. Mortality remained high even if larvae were switched to the dinoflagellate diet. Aldehydes, cleaved from a fatty acid precursor by enzymes activated within seconds after crushing of the cell, elicit the teratogenic effect. This insidious mechanism, which does not deter the herbivore from feeding but impairs its recruitment, will restrain the cohort size of the next generation of early-rising overwinterers. Such a transgenerational plant–herbivore interaction could explain the recurringly inefficient use of a predictable, potentially valuable food resource—the spring diatom bloom—by marine zooplankton.

Transformation of Nonselectable Reporter Genes in Marine Diatoms.

Abstract: We report the genetic transformation of two marine diatoms by microparticle bombardment. The pennate diatom Phaeodactylum tricornutum was transformed with the bacterial gene Sh ble from Streptoalloteichus hindustanus, which confers resistance to the antibiotics phleomycin and zeocin. Transformants contained between 1 and 10 copies of the exogenous DNA integrated into the genome by illegitimate recombination at apparently random locations. Transformation efficiencies were around 10−6, and individual cell lines could be maintained at −80°C following cryopreservation. Also, P. tricornutum could be transformed simultaneously with two different plasmids, one containing the Sh ble gene and another containing the firefly luciferase gene (LUC) under control of a promoter derived from a fucoxanthin, chlorophyll a/c-binding protein gene (FCP). In these cotransformants, LUC activity was light inducible. The transient transformation of the centric diatom Thalassiosira weissflogii with the bacterial β-glucuronidase (GUS) gene has also been achieved using similar transformation technology. The availability of gene transfer protocols for marine diatoms, together with a range of functional reporter genes and regulated expression systems, will permit molecular dissection of their biology and allow an assessment of the biotechnological potential of these organisms.

GROWTH INHIBITION AND TOXICITY OF THE DIATOM ALDEHYDE 2‐TRANS, 4‐TRANS‐DECADIENAL ON THALASSIOSIRA WEISSFLOGII (BACILLARIOPHYCEAE)

A common aldehyde present in marine and freshwater diatoms, 2‐trans, 4‐trans‐decadienal (A3), is involved in the wound‐activated response of diatoms to copepod grazing. Upon breakage of the diatom cell membrane, aldehydes are enzymatically produced by the rapid conversion of precursors and strongly impact copepod reproduction by impairing egg production and hatching success, inducing teratogenic embryos modifications. In this study, A3 was assayed with the marine diatom Thalassiosira weissflogii (Grunow) Fryxell et Hasle. The aldehyde concentration necessary to reduce 50% growth rate (EC50) was 0.29 mg·L−1. Decadienal was found to inhibit T. weissflogii cell growth in a dose‐ and time‐dependent manner, with irreversible effects after 24 h of exposure. Decadienal induced a degenerative process, through modifications of cell membrane characteristics, interference with cell cycle progression, and with cell metabolic activity, leading to cell death. A preferential action of A3 on dividing cells was observed. Photosynthetic efficiency significantly decreased upon exposure to the aldehyde, paralleled by an increase in diatoxanthin, suggesting a protective role of this xanthophyll, usually involved in photoprotection. Dying cells exhibited the morphological and biochemical features that bear close resemblance to apoptosis of mammalian cells, including cell shrinkage, chromatin condensation, and degradation of nuclear DNA to nucleosomal size fragments. These data are the first direct evidence to show aldehydes are toxic to diatoms. We suggest a possible nontoxic role of such compounds as chemical signals of unfavorable conditions within the phytoplankton communities, which may be relevant for the population dynamics of diatoms during blooms.

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.

Cell Cycle Regulation by Light in Prochlorococcus Strains

ABSTRACT The effect of light on the synchronization of cell cycling was investigated in several strains of the oceanic photosynthetic prokaryote Prochlorococcus using flow cytometry. When exposed to a light-dark (L-D) cycle with an irradiance of 25 μmol of quanta · m−2 s−1, the low-light-adapted strain SS 120 appeared to be better synchronized than the high-light-adapted strain PCC 9511. Submitting L-D-entrained populations to shifts (advances or delays) in the timing of the “light on” signal translated to corresponding shifts in the initiation of the S phase, suggesting that this signal is a key parameter for the synchronization of population cell cycles. Cultures that were shifted from an L-D cycle to continuous irradiance showed persistent diel oscillations of flow-cytometric signals (light scatter and chlorophyll fluorescence) but with significantly reduced amplitudes and a phase shift. Complete darkness arrested most of the cells in the G1 phase of the cell cycle, indicating that light is required to trigger the initiation of DNA replication and cell division. However, some cells also arrested in the S phase, suggesting that cell cycle controls in Prochlorococcus spp. are not as strict as in marine Synechococcus spp. ShiftingProchlorococcus cells from low to high irradiance translated quasi-instantaneously into an increase of cells in both the S and G2 phases of the cell cycle and then into faster growth, whereas the inverse shift induced rapid slowing of the population growth rate. These data suggest a close coupling between irradiance levels and cell cycling in Prochlorococcus spp.

Environmental characteristics of Agulhas rings affect interocean plankton transport

Agulhas rings provide the principal route for ocean waters to circulate from the Indo-Pacific to the Atlantic basin. Their influence on global ocean circulation is well known, but their role in plankton transport is largely unexplored. We show that, although the coarse taxonomic structure of plankton communities is continuous across the Agulhas choke point, South Atlantic plankton diversity is altered compared with Indian Ocean source populations. Modeling and in situ sampling of a young Agulhas ring indicate that strong vertical mixing drives complex nitrogen cycling, shaping community metabolism and biogeochemical signatures as the ring and associated plankton transit westward. The peculiar local environment inside Agulhas rings may provide a selective mechanism contributing to the limited dispersal of Indian Ocean plankton populations into the Atlantic.

Quantification of Dissolved and Particulate Polyunsaturated Aldehydes in the Adriatic Sea

Polyunsaturated aldehydes (PUA) are supposed to play critical roles in chemically-mediated plankton interactions. Laboratory studies suggest that they act as mediators of chemical defense and chemical communication. PUA are oxylipins containing an α,β,γ,δ–unsaturated aldehyde structure element and are mainly found in diatoms. We present here a detailed surface mapping of PUA during a spring bloom of the diatom Skeletonema marinoi in the Adriatic Sea. We monitored dissolved PUA, as well as particulate PUA, which are produced by phytoplankton after cell disintegration. Our survey revealed a patchy distribution of PUA and shows that at most stations S. marinoi is the major contributor to the overall PUA. Our data also suggest that lysis of a diatom bloom can contribute significantly to the dissolved PUA concentrations and that other producers, which are smaller in cell size compared to diatoms, have to be taken into account as well if the total PUA content of marine samples is considered. The analyses of samples collected in deeper water suggests that diatom contribution to PUA decreases with depth, while smaller-sized unidentified organisms take place as dominant contributors to the PUA concentrations.

High plasticity in the production of diatom-derived polyunsaturated aldehydes under nutrient limitation: physiological and ecological implications.

Diatoms have evolved a silicified cell wall that provides an efficient barrier against herbivores. These microalgae also produce chemical compounds such as polyunsaturated aldehydes (PUAs) that can potentially impair recruitment and cause malformations in the offspring of such grazers. We measured silica content as an indication of cell wall thickness, organic nutrient cell quotas, PUAs and polyunsaturated fatty acid cell content in Skeletonema marinoi grown under N-, P- and Si-limitation in continuous cultures. A 7.5 fold increase in PUA production was observed (27.5 fmol cell(-1)) in Si-limited cells with respect to the controls, while Si content decreased by 50%. PUA production decreased in nitrogen-limited cells to 0.14 fmol cell(-1) and increased 3 times in phosphorus-limited cells (7.53 fmol cell(-1)), while silica content increased by 20% in both cases. The substrate to product ratio suggested that production of the PUAs heptadienal and octadienal was limited by the amount of substrate under P- and Si-limitation, and by enzyme activity under N-limitation. Octatrienal production was likely limited by enzyme activity in all growth conditions. The high PUA levels produced under Si-limitation, when cells have a thinner cell wall, suggests a compensatory alternation of mechanical and chemical defense mechanisms.

Phytoplankton cell lysis associated with polyunsaturated aldehyde release in the Northern Adriatic Sea.

Diatoms are able to react to biotic and abiotic stress, such as competition, predation and unfavorable growth conditions, by producing bioactive compounds including polyunsaturated aldehydes (PUAs). PUAs have been shown to act against grazers and either enhance or inhibit the growth of different phytoplankton and bacteria both in culture and in the field. Presence of nanomolar concentrations of dissolved PUAs in seawater has been reported in the North Adriatic Sea (Mediterranean), suggesting that these compounds are released in seawater following diatom cell lysis. However, the origin of the PUAs and their effects on natural phytoplankton assemblages remain unclear. Here we present data from four oceanographic cruises that took place during diatom blooms in the northern Adriatic Sea where concentrations of particulate and dissolved PUAs were monitored along with phytoplankton cell lysis. Cell lysis was positively correlated with both concentrations of particulate and dissolved PUAs (R = 0.69 and R = 0.77, respectively), supporting the hypothesis that these compounds are released by cell lysis. However, the highest concentration of dissolved PUAs (2.53 nM) was measured when cell lysis was high (0.24 d−1) but no known PUA-producing diatoms were detected, suggesting either that other organisms can produce PUAs or that PUA-producing enzymes retain activity extracellularly after diatom cells have lysed. Although in situ concentrations of dissolved PUAs were one to three orders of magnitude lower than those typically used in laboratory culture experiments, we argue that concentrations produced in the field could induce similar effects to those observed in culture and therefore may help shape plankton community composition and function in the oceans.

Finding a partner in the ocean: molecular and evolutionary bases of the response to sexual cues in a planktonic diatom

Summary Microalgae play a major role as primary producers in aquatic ecosystems. Cell signalling regulates their interactions with the environment and other organisms, yet this process in phytoplankton is poorly defined. Using the marine planktonic diatom Pseudo‐nitzschia multistriata, we investigated the cell response to cues released during sexual reproduction, an event that demands strong regulatory mechanisms and impacts on population dynamics. We sequenced the genome of P. multistriata and performed phylogenomic and transcriptomic analyses, which allowed the definition of gene gains and losses, horizontal gene transfers, conservation and evolutionary rate of sex‐related genes. We also identified a small number of conserved noncoding elements. Sexual reproduction impacted on cell cycle progression and induced an asymmetric response of the opposite mating types. G protein‐coupled receptors and cyclic guanosine monophosphate (cGMP) are implicated in the response to sexual cues, which overall entails a modulation of cell cycle, meiosis‐related and nutrient transporter genes, suggesting a fine control of nutrient uptake even under nutrient‐replete conditions. The controllable life cycle and the genome sequence of P. multistriata allow the reconstruction of changes occurring in diatoms in a key phase of their life cycle, providing hints on the evolution and putative function of their genes and empowering studies on sexual reproduction.