Coastal phytoplankton blooms expand and intensify in the 21st century.

Phytoplankton blooms in coastal oceans can be beneficial to coastal fisheries production and ecosystem function, but can also cause major environmental problems1,2-yet detailed characterizations of bloom incidence and distribution are not available worldwide. Here we map daily marine coastal algal blooms between 2003 and 2020 using global satellite observations at 1-km spatial resolution. We found that algal blooms occurred in 126 out of the 153 coastal countries examined. Globally, the spatial extent (+13.2%) and frequency (+59.2%) of blooms increased significantly (P < 0.05) over the study period, whereas blooms weakened in tropical and subtropical areas of the Northern Hemisphere. We documented the relationship between the bloom trends and ocean circulation, and identified the stimulatory effects of recent increases in sea surface temperature. Our compilation of daily mapped coastal phytoplankton blooms provides the basis for global assessments of bloom risks and benefits, and for the formulation or evaluation of management or policy actions.

Evidence for massive and recurrent toxic blooms of Alexandrium catenella in the Alaskan Arctic.

Among the organisms that spread into and flourish in Arctic waters with rising temperatures and sea ice loss are toxic algae, a group of harmful algal bloom species that produce potent biotoxins. Alexandrium catenella, a cyst-forming dinoflagellate that causes paralytic shellfish poisoning worldwide, has been a significant threat to human health in southeastern Alaska for centuries. It is known to be transported into Arctic regions in waters transiting northward through the Bering Strait, yet there is little recognition of this organism as a human health concern north of the Strait. Here, we describe an exceptionally large A. catenella benthic cyst bed and hydrographic conditions across the Chukchi Sea that support germination and development of recurrent, locally originating and self-seeding blooms. Two prominent cyst accumulation zones result from deposition promoted by weak circulation. Cyst concentrations are among the highest reported globally for this species, and the cyst bed is at least 6× larger in area than any other. These extraordinary accumulations are attributed to repeated inputs from advected southern blooms and to localized cyst formation and deposition. Over the past two decades, warming has likely increased the magnitude of the germination flux twofold and advanced the timing of cell inoculation into the euphotic zone by 20 d. Conditions are also now favorable for bloom development in surface waters. The region is poised to support annually recurrent A. catenella blooms that are massive in scale, posing a significant and worrisome threat to public and ecosystem health in Alaskan Arctic communities where economies are subsistence based.

Insights into the loss factors of phytoplankton blooms: The role of cell mortality in the decline of two inshore Alexandrium blooms.

While considerable effort has been devoted to understanding the factors regulating the development of phytoplankton blooms, the mechanisms leading to bloom decline and termination have received less attention. Grazing and sedimentation have been invoked as the main routes for the loss of phytoplankton biomass, and more recently, viral lysis, parasitism and programmed cell death (PCD) have been recognized as additional removal factors. Despite the importance of bloom declines to phytoplankton dynamics, the incidence and significance of various loss factors in regulating phytoplankton populations have not been widely characterized in natural blooms. To understand mechanisms controlling bloom decline, we studied two independent, inshore blooms of Alexandrium fundyense, paying special attention to cell mortality as a loss pathway. We observed increases in the number of dead cells with PCD features after the peak of both blooms, demonstrating a role for cell mortality in their terminations. In both blooms, sexual cyst formation appears to have been the dominant process leading to bloom termination, as both blooms were dominated by small-sized gamete cells near their peaks. Cell death and parasitism became more significant as sources of cell loss several days after the onset of bloom decline. Our findings show two distinct phases of bloom decline, characterized by sexual fusion as the initial dominant cell removal processes followed by elimination of remaining cells by cell death and parasitism.

Bloom termination of the toxic dinoflagellate Alexandrium catenella: Vertical migration behavior, sediment infiltration, and benthic cyst yield.

New resting cyst production is crucial for the survival of many microbial eukaryotes including phytoplankton that cause harmful algal blooms. Production in situ has previously been estimated through sediment trap deployments, but here was instead assessed through estimation of the total number of planktonic cells and new resting cysts produced by a localized, inshore bloom of Alexandrium catenella, a dinoflagellate that is a globally important cause of paralytic shellfish poisoning. Our approach utilizes high frequency, automated water monitoring, weekly observation of new cyst production, and pre- and post-bloom spatial surveys of total resting cyst abundance. Through this approach, new cyst recruitment within the study area was shown to account for at least 10.9% ± 2.6% (SE) of the bloom's decline, ∼ 5× greater than reported from comparable, sediment trap based studies. The observed distribution and timing of new cyst recruitment indicate that: (1) planozygotes, the immediate precursor to cysts in the life cycle, migrate nearer to the water surface than other planktonic stages and (2) encystment occurs after planozygote settlement on bottom sediments. Near surface localization by planozygotes explains the ephemerality of red surface water discoloration by A. catenella blooms, and also enhances the dispersal of new cysts. Following settlement, bioturbation and perhaps active swimming promote sediment infiltration by planozygotes, reducing the extent of cyst redistribution between blooms. The concerted nature of bloom sexual induction, especially in the context of an observed upper limit to A. catenella bloom intensities and heightened susceptibility of planozygotes to the parasite Amoebophrya, is also discussed.

Effects of temperature and salinity on the growth of Alexandrium (Dinophyceae) isolates from the Salish Sea.

Toxin-producing blooms of dinoflagellates in the genus Alexandrium have plagued the inhabitants of the Salish Sea for centuries. Yet the environmental conditions that promote accelerated growth of this organism, a producer of paralytic shellfish toxins, is lacking. This study quantitatively determined the growth response of two Alexandrium isolates to a range of temperatures and salinities, factors that will strongly respond to future climate change scenarios. An empirical equation, derived from observed growth rates describing the temperature and salinity dependence of growth, was used to hindcast bloom risk. Hindcasting was achieved by comparing predicted growth rates, calculated from in situ temperature and salinity data from Quartermaster Harbor, with corresponding Alexandrium cell counts and shellfish toxin data. The greatest bloom risk, defined at µ >0.25 d(-1) , generally occurred from April through November annually; however, growth rates rarely fell below 0.10 d(-1) . Except for a few occasions, Alexandrium cells were only observed during the periods of highest bloom risk and paralytic shellfish toxins above the regulatory limit always fell within the periods of predicted bloom occurrence. While acknowledging that Alexandrium growth rates are affected by other abiotic and biotic factors, such as grazing pressure and nutrient availability, the use of this empirical growth function to predict higher risk time frames for blooms and toxic shellfish within the Salish Sea provides the groundwork for a more comprehensive biological model of Alexandrium bloom dynamics in the region and will enhance our ability to forecast blooms in the Salish Sea under future climate change scenarios.

The effect of the toxic dinoflagellate Alexandrium fundyense on the fitness of the calanoid copepod Calanus finmarchicus.

Inshore and offshore waters of the Gulf of Maine (USA) have spring/summer harmful algal blooms (HABs) of the toxic dinoflagellate Alexandrium fundyense, which is responsible for paralytic shellfish poisoning (PSP) in humans. The calanoid copepod Calanus finmarchicus co-occurs with A. fundyense during the seasonal blooms. At that time, C. finmarchicus population abundances are high, dominated by immature copepods preparing for diapause, and by actively-reproducing adults. High survival has been reported for copepods exposed to toxic A. fundyense, but little is known about possible sublethal effects. In this study, C. finmarchicus adult females were fed either a control diet of non-toxic Rhodomonas spp. or one of two diets containing either low dose (LD) or high dose (HD) levels (50 and 200 cells mL-1, respectively) of toxic A. fundyense for a total of 7 days in two independent experiments. As expected, ingestion of the dinoflagellate had no effect on copepod survival and grazing activity. However, significant reductions of egg production and egg viability were observed in C. finmarchicus females fed on either experimental diet. After the 7-day experiment, total nauplius production by females on the LD and HD diets was reduced by 35% to 75% compared to the control females. These results suggest that blooms of A. fundyense in the Gulf of Maine may be an environmental challenge for C. finmarchicus populations, with a potential negative effect on copepod recruitment.

Rapid growth and concerted sexual transitions by a bloom of the harmful dinoflagellate Alexandrium fundyense (Dinophyceae).

Transitions between life cycle stages by the harmful dinoflagellate Alexandrium fundyense are critical for the initiation and termination of its blooms. To quantify these transitions in a single population, an Imaging FlowCytobot (IFCB), was deployed in Salt Pond (Eastham, Massachusetts), a small, tidally flushed kettle pond that hosts near annual, localized A. fundyense blooms. Machine-based image classifiers differentiating A. fundyense life cycle stages were developed and results were compared to manually corrected IFCB samples, manual microscopy-based estimates of A. fundyense abundance, previously published data describing prevalence of the parasite Amoebophrya, and a continuous culture of A. fundyense infected with Amoebophrya. In Salt Pond, a development phase of sustained vegetative division lasted approximately 3 weeks and was followed by a rapid and near complete conversion to small, gamete cells. The gametic period (∼3 d) coincided with a spike in the frequency of fusing gametes (up to 5% of A. fundyense images) and was followed by a zygotic phase (∼4 d) during which cell sizes returned to their normal range but cell division and diel vertical migration ceased. Cell division during bloom development was strongly phased, enabling estimation of daily rates of division, which were more than twice those predicted from batch cultures grown at similar temperatures in replete medium. Data from the Salt Pond deployment provide the first continuous record of an A. fundyense population through its complete bloom cycle and demonstrate growth and sexual induction rates much higher than are typically observed in culture.

The effects of elevated CO2 on the growth and toxicity of field populations and cultures of the saxitoxin-producing dinoflagellate, Alexandrium fundyense.

The effects of coastal acidification on the growth and toxicity of the saxitoxin-producing dinoflagellate Alexandrium fundyense were examined in culture and ecosystem studies. In culture experiments, Alexandrium strains isolated from Northport Bay NY, USA, and the Bay of Fundy, Canada, grew significantly faster (16 -190%; p<0.05) when exposed to elevated levels of pCO2 (~ 800- 1900µatm) compared to lower levels (~390µatm). Exposure to higher levels of pCO2 also resulted in significant increases (71 - 81%) in total cellular toxicity (fg STX eq. cell-1) in the Northport Bay strain, while no changes in toxicity were detected in the Bay of Fundy strain. The positive relationship between pCO2 enhancement and elevated growth was reproducible using natural populations from Northport; Alexandrium densities were significantly and consistently enhanced when natural populations were incubated at 1500 µatm pCO2, a value at the upper range of those recorded in Northport Bay, 390 - 1500 µatm. During natural Alexandrium blooms in Northport Bay, pCO2 concentrations increased over the course of a bloom to more than 1700µatm and were highest in regions with the greatest Alexandrium abundances, suggesting Alexandrium may be further exacerbating acidification or be especially adapted to these extreme, acidified conditions. The co-occurrence of Alexandrium blooms and elevated pCO2 represents a previously unrecognized, compounding environmental threat to coastal ecosystems. The ability of elevated pCO2 to enhance the growth and toxicity of Alexandrium indicates that acidification promoted by eutrophication or climate change can intensify these, and perhaps other, harmful algal blooms.

Characterization and comparison of toxin-producing isolates of Dinophysis acuminata from New England and Canada.

Following the identification of the first toxic isolate of Dinophysis acuminata from the northwestern Atlantic, we conducted detailed investigations into the morphology, phylogeny, physiology, and toxigenicity of three isolates from three sites within the northeastern U.S./Canada region: Eel Pond and Martha's Vineyard, Massachusetts, and the Bay of Fundy. Another isolate, collected from the Gulf of Mexico, was grown under the same light, temperature, and prey conditions for comparison. Despite observed phenotypic heterogeneity, morphometrics and molecular evidence classified the three northwestern Atlantic isolates as D. acuminata Claparède & Lachmann, whereas the isolate from the Gulf of Mexico was morphologically identified as D. cf. ovum. Physiological and toxin analyses supported these classifications, with the three northwestern Atlantic isolates being more similar to each other with respect to growth rate, toxin profile, and diarrhetic shellfish poisoning (DSP) toxin content (okadaic acid + dinophysistoxin 1/cell) than they were to the isolate from the Gulf of Mexico, which had toxin profiles similar to those published for D. cf. ovum F. Schütt. The DSP toxin content, 0.01-1.8 pg okadaic acid (OA) + dinophysistoxin (DTX1) per cell, of the three northwestern Atlantic isolates was low relative to other D. acuminata strains from elsewhere in the world, consistent with the relative scarcity of shellfish harvesting closures due to DSP toxins in the northeastern U.S. and Canada. If this pattern is repeated with the analyses of more geographically and temporally dispersed isolates from the region, it would appear that the risk of significant DSP toxin outbreaks in the northwestern Atlantic is low to moderate. Finally, the morphological, physiological, and toxicological variability within D. acuminata may reflect spatial (and/or temporal) population structure, and suggests that sub-specific resolution may be helpful in characterizing bloom dynamics and predicting toxicity.

Zooplankton Community Grazing Impact on a Toxic Bloom of Alexandrium fundyense in the Nauset Marsh System, Cape Cod, Massachusetts, USA.

Embayments and salt ponds along the coast of Massachusetts can host localized blooms of the toxic dinoflagellate Alexandrium fundyense. One such system, exhibiting a long history of toxicity and annual closures of shellfish beds, is the Nauset Marsh System (NMS) on Cape Cod. In order measure net growth rates of natural A. fundyense populations in the NMS during spring 2012, incubation experiments were conducted on seawater samples from two salt ponds within the NMS (Salt Pond and Mill Pond). Seawater samples containing natural populations of grazers and A. fundyense were incubated at ambient temperatures. Concentrations of A. fundyense after incubations were compared to initial abundances to determine net increases from population growth, or decreases presumed to be primarily due to grazing losses. Abundances of both microzooplankton (ciliates, rotifers, copepod nauplii and heterotrophic dinoflagellates) and mesozooplankton (copepodites and adult copepods, marine cladocerans, and meroplankton) grazers were also determined. This study documented net growth rates that were highly variable throughout the bloom, calculated from weekly bloom cell counts from the start of sampling to bloom peak in both ponds (Mill Pond range = 0.12 - 0.46 d-1; Salt Pond range = -0.02 - 0.44 d-1). Microzooplankton grazers that were observed with ingested A. fundyense cells included polychaete larvae, rotifers, tintinnids, and heterotrophic dinoflagellates of the genera Polykrikos and Gymnodinium. Significant A. fundyense net growth was observed in two incubation experiments, and only a single experiment exhibited significant population losses. For the majority of experiments, due to high variability in data, net changes in A. fundyense abundance were not significant after the 24-hr incubations. However, experimental net growth rates through bloom peak were not statistically distinguishable from estimated long-term average net growth rates of natural populations in each pond (Mill Pond = 0.27 d-1 and Salt Pond = 0.20 d-1), which led to peak bloom concentrations on the order of 106 cells l-1 in both ponds. Experimental net growth rates from the incubations underestimated the observed natural net growth rates at several time intervals prior to bloom peak, which may indicate that natural populations experienced additional sources of vegetative cells or periods of reduced losses that the 24-hr incubation experiments did not capture, or that the experimental procedure introduced containment artifacts.

Factors regulating excystment of Alexandrium in Puget Sound, WA, USA.

Factors regulating excystment of a toxic dinoflagellate in the genus Alexandrium were investigated in cysts from Puget Sound, Washington State, USA. Experiments were carried out in the laboratory using cysts collected from benthic seedbeds to determine if excystment is controlled by internal or environmental factors. The results suggest that the timing of germination is not tightly controlled by an endogenous clock, though there is a suggestion of a cyclical pattern. This was explored using cysts that had been stored under cold (4 °C), anoxic conditions in the dark and then incubated for 6 weeks at constant favorable environmental conditions. Excystment occurred during all months of the year, with variable excystment success ranging from 31-90%. When cysts were isolated directly from freshly collected sediments every month and incubated at the in situ bottom water temperature, a seasonal pattern in excystment was observed that was independent of temperature. This pattern may be consistent with secondary dormancy, an externally modulated pattern that prevents excystment during periods that are not favorable for sustained vegetative growth. However, observation over more annual cycles is required and the duration of the mandatory dormancy period of these cysts must be determined before the seasonality of germination can be fully characterized in Alexandrium from Puget Sound. Both temperature and light were found to be important environmental factors regulating excystment, with the highest rates of excystment observed for the warmest temperature treatment (20 °C) and in the light.

Harmful algal blooms and climate change: Learning from the past and present to forecast the future.

Climate change pressures will influence marine planktonic systems globally, and it is conceivable that harmful algal blooms may increase in frequency and severity. These pressures will be manifest as alterations in temperature, stratification, light, ocean acidification, precipitation-induced nutrient inputs, and grazing, but absence of fundamental knowledge of the mechanisms driving harmful algal blooms frustrates most hope of forecasting their future prevalence. Summarized here is the consensus of a recent workshop held to address what currently is known and not known about the environmental conditions that favor initiation and maintenance of harmful algal blooms. There is expectation that harmful algal bloom (HAB) geographical domains should expand in some cases, as will seasonal windows of opportunity for harmful algal blooms at higher latitudes. Nonetheless there is only basic information to speculate upon which regions or habitats HAB species may be the most resilient or susceptible. Moreover, current research strategies are not well suited to inform these fundamental linkages. There is a critical absence of tenable hypotheses for how climate pressures mechanistically affect HAB species, and the lack of uniform experimental protocols limits the quantitative cross-investigation comparisons essential to advancement. A HAB "best practices" manual would help foster more uniform research strategies and protocols, and selection of a small target list of model HAB species or isolates for study would greatly promote the accumulation of knowledge. Despite the need to focus on keystone species, more studies need to address strain variability within species, their responses under multifactorial conditions, and the retrospective analyses of long-term plankton and cyst core data; research topics that are departures from the norm. Examples of some fundamental unknowns include how larger and more frequent extreme weather events may break down natural biogeographic barriers, how stratification may enhance or diminish HAB events, how trace nutrients (metals, vitamins) influence cell toxicity, and how grazing pressures may leverage, or mitigate HAB development. There is an absence of high quality time-series data in most regions currently experiencing HAB outbreaks, and little if any data from regions expected to develop HAB events in the future. A subset of observer sites is recommended to help develop stronger linkages among global, national, and regional climate change and HAB observation programs, providing fundamental datasets for investigating global changes in the prevalence of harmful algal blooms. Forecasting changes in HAB patterns over the next few decades will depend critically upon considering harmful algal blooms within the competitive context of plankton communities, and linking these insights to ecosystem, oceanographic and climate models. From a broader perspective, the nexus of HAB science and the social sciences of harmful algal blooms is inadequate and prevents quantitative assessment of impacts of future HAB changes on human well-being. These and other fundamental changes in HAB research will be necessary if HAB science is to obtain compelling evidence that climate change has caused alterations in HAB distributions, prevalence or character, and to develop the theoretical, experimental, and empirical evidence explaining the mechanisms underpinning these ecological shifts.

Cellular responses associated with ROS production and cell fate decision in early stress response to iron limitation in the diatom Thalassiosira pseudonana.

Investigation of how diatoms cope with the rapid fluctuations in iron bioavailability in marine environments may facilitate a better understanding of the mechanisms underlying their ecological success, in particular their ability to proliferate rapidly during favorable conditions. In this study, using in vivo biochemical markers and whole-cell iTRAQ-based proteomics analysis, we explored the cellular responses associated with reactive oxygen species (ROS) production and cell fate decision during the early response to Fe limitation in the centric diatom Thalassiosira pseudonana. Fe limitation caused a significant decrease in Photosystem (PS) II photosynthetic efficiency, damage to the photosynthetic electron transport chain in PS I, and blockage of the respiratory chain in complexes III and IV, which could all result in excess ROS accumulation. The increase in ROS likely triggered programmed cell death (PCD) in some of the Fe-limited cells through synthesis of a series of proteins involved in the delicate balance between pro-survival and pro-PCD factors. The results provide molecular-level insights into the major strategies that may be employed by T. pseudonana in response to Fe-limitation: the reduction of cell population density through PCD to reduce competition for available Fe, the reallocation of intracellular nitrogen and Fe to ensure survival, and an increase in expression of antioxidant and anti-PCD proteins to cope with stress.

Evolution of saxitoxin synthesis in cyanobacteria and dinoflagellates.

Dinoflagellates produce a variety of toxic secondary metabolites that have a significant impact on marine ecosystems and fisheries. Saxitoxin (STX), the cause of paralytic shellfish poisoning, is produced by three marine dinoflagellate genera and is also made by some freshwater cyanobacteria. Genes involved in STX synthesis have been identified in cyanobacteria but are yet to be reported in the massive genomes of dinoflagellates. We have assembled comprehensive transcriptome data sets for several STX-producing dinoflagellates and a related non-toxic species and have identified 265 putative homologs of 13 cyanobacterial STX synthesis genes, including all of the genes directly involved in toxin synthesis. Putative homologs of four proteins group closely in phylogenies with cyanobacteria and are likely the functional homologs of sxtA, sxtG, and sxtB in dinoflagellates. However, the phylogenies do not support the transfer of these genes directly between toxic cyanobacteria and dinoflagellates. SxtA is split into two proteins in the dinoflagellates corresponding to the N-terminal portion containing the methyltransferase and acyl carrier protein domains and a C-terminal portion with the aminotransferase domain. Homologs of sxtB and N-terminal sxtA are present in non-toxic strains, suggesting their functions may not be limited to saxitoxin production. Only homologs of the C-terminus of sxtA and sxtG were found exclusively in toxic strains. A more thorough survey of STX+ dinoflagellates will be needed to determine if these two genes may be specific to SXT production in dinoflagellates. The A. tamarense transcriptome does not contain homologs for the remaining STX genes. Nevertheless, we identified candidate genes with similar predicted biochemical activities that account for the missing functions. These results suggest that the STX synthesis pathway was likely assembled independently in the distantly related cyanobacteria and dinoflagellates, although using some evolutionarily related proteins. The biological role of STX is not well understood in either cyanobacteria or dinoflagellates. However, STX production in these two ecologically distinct groups of organisms suggests that this toxin confers a benefit to producers that we do not yet fully understand.

Temperature dependence of an estuarine harmful algal bloom: Resolving interannual variability in bloom dynamics using a degree day approach.

Observations of harmful algal blooms (HABs) of the dinoflagellate Alexandrium fundyense in an estuary over multiple years were used to assess drivers of their spatial and temporal variability. Nauset Estuary on Cape Cod, Massachusetts has a recurrent, self-seeding A. fundyense population that produces paralytic shellfish poisoning toxins and leads to nearly annual closure to shellfishing. Weekly surveys of the entire estuary were made in 3 of 4 consecutive years, with surveys of a subembayment during the intervening year. Major A. fundyense blooms were observed all 4 years, with maximum concentrations >106 cells L-1. Concentrations were greatest in three salt ponds at the distal edges of the estuary. The bloom timing varied among the salt ponds and among years, although the blooms had similar durations and maximum cell concentrations. Nutrient concentrations did not correlate with the growth of the bloom, but differences in water temperature among years and ponds were significant. Net growth rates inferred from the surveys were similar to those from laboratory experiments, and increased linearly with temperature. A growing degree day calculation was used to account for effects of interannual variability and spatial gradients in water temperature on population development. The approach collapsed variability in the timing of bloom onset, development, and termination across years and among ponds, suggesting that this relatively simple metric could be used as an early-warning indicator for HABs in Nauset and similar areas with localized, self-seeding blooms.

Alexandrium fundyense cyst viability and germling survival in light vs. dark at a constant low temperature.

Both observations and models suggest that large-scale coastal blooms of Alexandrium fundyense in the Gulf of Maine are seeded by deep-bottom cyst accumulation zones ("seed beds") where cysts germinate from the sediment surface or the overlying near-bottom nepheloid layers at water depths exceeding 100 m. The germling cells and their vegetative progeny are assumed to be subject to mortality while in complete darkness, as they swim to illuminated surface waters. To test the validity of this assumption we conducted laboratory investigations of cyst viability and the survival of the germling cells and their vegetative progeny during prolonged exposure to darkness at a temperature of 6 °C, simulating the conditions in deep Gulf of Maine waters. We isolated cysts from bottom sediments collected in the Gulf of Maine under low red light and incubated them in 96-well tissue culture-plates in culture medium under a 10:14 h light:dark cycle and under complete darkness. Cyst viability was high, with excystment frequency reaching 90% in the illuminated treatment after 30 days and in the dark treatment after 50 days. Average germination rates were 0.062 and 0.038 d-1 for light and dark treatments, respectively. The dark treatment showed an approximately 2-week time lag in maximum germination rates compared to the light treatment. Survival of germlings was considerably lower in the dark treatment. In the light treatments, 47% of germinated cysts produced germlings that were able to survive for 7 days and produce vegetative progeny, i.e., there were live cells in the well along with an empty cyst at least once during the experiment. In the dark treatments 12% of the cysts produced germlings that were able to survive for the same length of time. When dark treatments are scaled to take into account non-darkness related mortality, approximately 28% of the cysts produced germlings that were able to survive for at least 7 days. Even though cysts are able to germinate in darkness, the lack of illumination considerably reduces survival rate of germling cells. In addition to viability of cysts in surface sediments and the near-bottom nepheloid layer, survivability of germling cells and their vegetative progeny at aphotic depths is an important consideration in assessing the quantitative role of deep-coastal cyst seed beds in bloom formation.

Complexities of bloom dynamics in the toxic dinoflagellate Alexandrium fundyense revealed through DNA measurements by imaging flow cytometry coupled with species-specific rRNA probes.

Measurements of the DNA content of different protist populations can shed light on a variety of processes, including cell division, sex, prey ingestion, and parasite invasion. Here, we modified an Imaging FlowCytobot (IFCB), a custom-built flow cytometer that records images of microplankton, to measure the DNA content of large dinoflagellates and other high-DNA content species. The IFCB was also configured to measure fluorescence from Cy3-labeled rRNA probes, aiding the identification of Alexandrium fundyense (syn. A. tamarense Group I), a photosynthetic dinoflagellate that causes paralytic shellfish poisoning (PSP). The modified IFCB was used to analyze samples from the development, peak and termination phases of an inshore A. fundyense bloom (Salt Pond, Eastham, MA USA), and from a rare A. fundyense 'red tide' that occurred in the western Gulf of Maine, offshore of Portsmouth, NH (USA). Diploid or G2 phase ('2C') A. fundyense cells were frequently enriched at the near-surface, suggesting an important role for aggregation at the air-sea interface during sexual events. Also, our analysis showed that large proportions of A. fundyense cells in both the Salt Pond and red tide blooms were planozygotes during bloom decline, highlighting the importance of sexual fusion to bloom termination. At Salt Pond, bloom decline also coincided with a dramatic rise in infections by the parasite genus Amoebophrya. The samples that were most heavily infected contained many large cells with higher DNA-associated fluorescence than 2C vegetative cells, but these cells' nuclei were also frequently consumed by Amoebophrya trophonts. Neither large cell size nor increased DNA-associated fluorescence could be replicated by infecting an A. fundyense culture of vegetative cells. Therefore we attribute these characteristics of the large Salt Pond cells to planozygote maturation rather than Amoebophrya infection, though an interaction between infection and planozygote maturation may also have contributed. The modified IFCB is a valuable tool for exploring the conditions that promote sexual transitions by dinoflagellate blooms but care is needed when interpreting results from samples in which parasitism is prevalent.

Investigating the importance of sediment resuspension in Alexandrium fundyense cyst population dynamics in the Gulf of Maine.

Cysts of Alexandrium fundyense, a dinoflagellate that causes toxic algal blooms in the Gulf of Maine, spend the winter as dormant cells in the upper layer of bottom sediment or the bottom nepheloid layer and germinate in spring to initiate new blooms. Erosion measurements were made on sediment cores collected at seven stations in the Gulf of Maine in the autumn of 2011 to explore if resuspension (by waves and currents) could change the distribution of over-wintering cysts from patterns observed in the previous autumn; or if resuspension could contribute cysts to the water column during spring when cysts are viable. The mass of sediment eroded from the core surface at 0.4 Pa ranged from 0.05 kg m-2 near Grand Manan Island, to 0.35 kg m-2 in northern Wilkinson Basin. The depth of sediment eroded ranged from about 0.05 mm at a station with sandy sediment at 70 m water depth on the western Maine shelf, to about 1.2 mm in clayey-silt sediment at 250 m water depth in northern Wilkinson Basin. The sediment erodibility measurements were used in a sediment-transport model forced with modeled waves and currents for the period October 1, 2010 to May 31, 2011 to predict resuspension and bed erosion. The simulated spatial distribution and variation of bottom shear stress was controlled by the strength of the semi-diurnal tidal currents, which decrease from east to west along the Maine coast, and oscillatory wave-induced currents, which are strongest in shallow water. Simulations showed occasional sediment resuspension along the central and western Maine coast associated with storms, steady resuspension on the eastern Maine shelf and in the Bay of Fundy associated with tidal currents, no resuspension in northern Wilkinson Basin, and very small resuspension in western Jordan Basin. The sediment response in the model depended primarily on the profile of sediment erodibility, strength and time history of bottom stress, consolidation time scale, and the current in the water column. Based on analysis of wave data from offshore buoys from 1996 to 2012, the number of wave events inducing a bottom shear stress large enough to resuspend sediment at 80 m ranged from 0 to 2 in spring (April and May) and 0 to 10 in winter (October through March). Wave-induced resuspension is unlikely in water greater than about 100 m deep. The observations and model results suggest that a millimeter or so of sediment and associated cysts may be mobilized in both winter and spring, and that the frequency of resuspension will vary interannually. Depending on cyst concentration in the sediment and the vertical distribution in the water column, these events could result in a concentration in the water column of at least 104 cysts m-3. In some years, resuspension events could episodically introduce cysts into the water column in spring, where germination is likely to be facilitated at the time of bloom formation. An assessment of the quantitative effects of cyst resuspension on bloom dynamics in any particular year requires more detailed investigation.

Categorizing the severity of paralytic shellfish poisoning outbreaks in the Gulf of Maine for forecasting and management.

Development of forecasting systems for harmful algal blooms (HABs) has been a long-standing research and management goal. Significant progress has been made in the Gulf of Maine, where seasonal bloom forecasts are now being issued annually using Alexandrium fundyense cyst abundance maps and a population dynamics model developed for that organism. Thus far, these forecasts have used terms such as "significant", "moderately large" or "moderate" to convey the extent of forecasted paralytic shellfish poisoning (PSP) outbreaks. In this study, historical shellfish harvesting closure data along the coast of the Gulf of Maine were used to derive a series of bloom severity levels that are analogous to those used to define major storms like hurricanes or tornados. Thirty-four years of PSP-related shellfish closure data for Maine, Massachusetts and New Hampshire were collected and mapped to depict the extent of coastline closure in each year. Due to fractal considerations, different methods were explored for measuring length of coastline closed. Ultimately, a simple procedure was developed using arbitrary straight-line segments to represent specific sections of the coastline. This method was consistently applied to each year's PSP toxicity closure map to calculate the total length of coastline closed. Maps were then clustered together statistically to yield distinct groups of years with similar characteristics. A series of categories or levels was defined ("Level 1: Limited", "Level 2: Moderate", and "Level 3: Extensive") each with an associated range of expected coastline closed, which can now be used instead of vague descriptors in future forecasts. This will provide scientifically consistent and simply defined information to the public as well as resource managers who make decisions on the basis of the forecasts.

PSP toxin levels and plankton community composition and abundance in size-fractionated vertical profiles during spring/summer blooms of the toxic dinoflagellate Alexandrium fundyense in the Gulf of Maine and on Georges Bank, 2007, 2008, and 2010: 2. Plankton community composition and abundance.

As part of the Gulf of Maine Toxicity (GOMTOX) project, we determined Alexandrium fundyense abundance, paralytic shellfish poisoning (PSP) toxin levels in various plankton size fractions, and the community composition of potential grazers of A. fundyense in plankton size fractions during blooms of this toxic dinoflagellate in the coastal Gulf of Maine and on Georges Bank in spring and summer of 2007, 2008, and 2010. PSP toxins and A. fundyense cells were found throughout the sampled water column (down to 50 m) in the 20-64 µm size fractions. While PSP toxins were widespread throughout all size classes of the zooplankton grazing community, the majority of the toxin was measured in the 20-64 µm size fraction. A. fundyense cellular toxin content estimated from field samples was significantly higher in the coastal Gulf of Maine than on Georges Bank. Most samples containing PSP toxins in the present study had diverse assemblages of grazers. However, some samples clearly suggested PSP toxin accumulation in several different grazer taxa including tintinnids, heterotrophic dinoflagellates of the genus Protoperidinium, barnacle nauplii, the harpacticoid copepod Microsetella norvegica, the calanoid copepods Calanus finmarchicus and Pseudocalanus spp., the marine cladoceran Evadne nordmanni, and hydroids of the genus Clytia. Thus, a diverse assemblage of zooplankton grazers accumulated PSP toxins through food-web interactions. This raises the question of whether PSP toxins pose a potential human health risk not only from nearshore bivalve shellfish, but also potentially from fish and other upper-level consumers in zooplankton-based pelagic food webs.