Expression of nuclear-encoded, haptophyte-derived ftsH genes support extremely rapid PSII repair and high-light photoacclimation in Karenia brevis (Dinophyceae).

Karenia brevis, a neurotoxic dinoflagellate that produces brevetoxins, is endemic to the Gulf of Mexico and can grow at high irradiances typical of surface waters found there. To build upon a growing number of studies addressing high-light tolerance in K. brevis, specific photobiology and molecular mechanisms underlying this capacity were evaluated in culture. Since photosystem II (PSII) repair cycle activity can be crucial to high light tolerance in plants and algae, the present study assessed this capacity in K. brevis and characterized the ftsH-like genes which are fundamental to this process. Compared with cultures grown in low-light, cultures grown in high-light showed a 65-fold increase in PSII photoinactivation, a ∼50-fold increase in PSII repair, enhanced nonphotochemical quenching (NPQ), and depressed Fv/Fm. Repair rates were among the fastest reported in phytoplankton. Publicly available K. brevis transcriptomes (MMETSP) were queried for ftsH-like sequences and refined with additional sequencing from two K. brevis strains. The genes were phylogenetically related to haptophyte orthologs, implicating acquisition during tertiary endosymbiosis. RT-qPCR of three of the four ftsH-like homologs revealed that poly-A tails predominated in all homologs, and that the most highly expressed homolog had a 5' splice leader and amino-acid motifs characteristic of chloroplast targeting, indicating nuclear encoding for this plastid-targeted gene. High-light cultures showed a ∼1.5-fold upregulation in mRNA expression of the thylakoid-associated genes. Overall, in conjunction with NPQ mechanisms, rapid PSII repair mediated by a haptophyte-derived ftsH prevents chronic photoinhibition in K. brevis. Our findings continue to build the case that high-light photobiology-supported by the acquisition and maintenance of tertiary endosymbiotic genes-is critical to the success of K. brevis in the Gulf of Mexico.

High-resolution Spatiotemporal Dynamics of Harmful Algae in the Indian River Lagoon (Florida)-A Case Study of Aureoumbra lagunensis, Pyrodinium bahamense, and Pseudo-nitzschia.

The Indian River Lagoon (IRL), located on the east coast of Florida, is a complex estuarine ecosystem that is negatively affected by recurring harmful algal blooms (HABs) from distinct taxonomic/functional groups. Enhanced monitoring was established to facilitate rapid quantification of three recurrent bloom taxa, Aureoumbra lagunensis, Pyrodinium bahamense, and Pseudo-nitzschia spp., and included corroborating techniques to improve the identification of small-celled nanoplankton (<10 µm in diameter). Identification and enumeration of these target taxa were conducted during 2015-2020 using a combination of light microscopy and species-specific approaches, specifically immunofluorescence flow cytometry as well as a newly developed qPCR assay for A. lagunensis presented here for the first time. An annual bloom index (ABI) was established for each taxon based on occurrence and abundance data. Blooms of A. lagunensis (>2×108 cells L-1) were observed in all six years sampled and across multiple seasons. In contrast, abundance of P. bahamense, largely driven by the annual temperature cycle that moderates life cycle transitions and growth, displayed a strong seasonal pattern with blooms (105-107 cells L-1) generally developing in early summer and subsiding in autumn. However, P. bahamense bloom development was delayed and abundance was significantly lower in years and locations with sustained A. lagunensis blooms. Pseudo-nitzschia spp. were broadly distributed with sporadic bloom concentrations (reaching 107 cells L-1), but with minimal concentrations of the toxin domoic acid detected (<0.02 µg L-1). In summer 2020, multiple monitoring tools characterized a novel nano-cyanobacterium bloom (reaching 109 cells L-1) that coincided with a decline in A. lagunensis and persisted into autumn. Statistical and time-series analyses of this spatiotemporally intensive dataset highlight prominent patterns in variability for some taxa, but also identifies challenges of characterizing mechanisms underlying more episodic yet persistent events. Nevertheless, the intersect of temperature and salinity as environmental proxies proved to be informative in delineating niche partitioning, not only in the case of taxa with long-standing data sets but also for seemingly unprecedented blooms of novel nanoplanktonic taxa.

Is Karenia brevis really a low-light-adapted species?

Despite nearly annual blooms of the neurotoxic dinoflagellate Karenia brevis (Davis) G. Hansen and Moestrup in the Gulf of Mexico, defining the suite of biological traits that explain its proliferation has remained challenging. Studies have described K. brevis as a low-light-adapted species, incapable of sustaining growth under high light, which is at odds with observed surface aggregations sometimes within centimeters of the sea surface and also with short-term experiments showing photosynthetic machinery accommodating high irradiances. Here, growth and photophysiology of three K. brevis isolates were evaluated under a range of environmentally relevant irradiances (10-1500 µmol photons m-2 s-1) in the laboratory. No differences in growth-irradiance curves were observed among isolates; all sustained maximum growth rates at the highest irradiances examined, even in exposures as long as three weeks. The growth efficiency α of K. brevis under light-limiting conditions appeared mediocre among dinoflagellates, and poorer than that of other phytoplankton (e.g., diatoms, cyanobacteria), implying that K. brevis is not a low-light specialist. This finding substantially alters earlier parameterizations of K. brevis growth-irradiance curves. Therefore, a model was developed to contextualize how these new growth-irradiance curves might affect bottom growth rates. This model was subsequently applied to a case study comparing seasonal light forcing offshore of Pinellas County, FL, USA, with a single empirical value for light attenuation, and seasonal bottom water temperatures. Predictions suggested that light may limit bottom growth as close as 1 km from shore in winter, but would only begin limiting growth 20 km from shore in summer. Population maintenance (no net growth) was possible as far offshore as 90 km in summer and 68 km in winter. These ranges intercept areas thought to be important for bloom initiation.

Cell cycle arrest and biochemical changes accompanying cell death in harmful dinoflagellates following exposure to bacterial algicide IRI-160AA.

Bacteria may play a role in regulating harmful algal blooms, but little is known about the biochemical and physiological changes associated with cell death induced by algicidal bacteria. Previous work characterized an algicidal exudate (IRI-160AA) produced by Shewanella sp. IRI-160 that is effective against dinoflagellates, while having little to no effect on other phytoplankton species in laboratory culture experiments. The objective of this study was to evaluate biochemical changes associated with cell death and impacts on the cell cycle in three dinoflagellate species (Prorocentrum minimum, Karlodinium veneficum and Gyrodinium instriatum) after exposure to IRI-160AA. In this study, IRI-160AA induced cell cycle arrest in all dinoflagellates examined. Several indicators for programmed cell death (PCD) that are often observed in phytoplankton in response to a variety of stressors were also evaluated. Cell death was accompanied by significant increases in DNA degradation, intra- and extracellular ROS concentrations and DEVDase (caspase-3 like) protease activity, which have been associated with PCD in other phytoplankton species. Overall, results of this investigation provide strong evidence that treatment with the bacterial algicide, IRI-160AA results in cell cycle arrest and induces biochemical changes consistent with stress-related cell death responses observed in other phytoplankton.

Effects of the bacterial algicide IRI-160AA on cellular morphology of harmful dinoflagellates.

The algicide, IRI-160AA, induces mortality in dinoflagellates but not other species of algae, suggesting that a shared characteristic or feature renders this class of phytoplankton vulnerable to the algicide. In contrast to other eukaryotic species, the genome of dinoflagellates is stabilized by high concentrations of divalent cations and transition metals and contains large amounts of DNA with unusual base modifications. These distinctions set dinoflagellates apart from other phytoplankton and suggest that the nucleus may be a dinoflagellate-specific target for IRI-160AA. In this study, morphological and ultrastructural changes in three dinoflagellate species, Prorocentrum minimum, Karlodinium veneficum and Gyrodinium instriatum, were evaluated after short-term exposure to IRI-160AA using super resolution structured illumination microscopy (SR-SIM) and transmission electron microscopy (TEM). Exposure to the algicide resulted in cytoplasmic membrane blebbing, differing chloroplast morphologies, nuclear expansion, and chromosome expulsion and/or destabilization. TEM analysis showed that chromosomes of algicide-treated K. veneficum appeared electron dense with fibrous protrusions. In algicide-treated P. minimum and G. instriatum, chromosome decompaction occurred, while for P. minimum, nuclear expulsion was also observed for several cells. Results of this investigation demonstrate that exposure to the algicide destabilizes dinoflagellate chromosomes, although it was not clear if the nucleus was the primary target of the algicide or if the observed effects on chromosomal structure were due to downstream impacts. In all cases, changes in cellular morphology and ultrastructure were observed within two hours, suggesting that the algicide may be an effective and rapid approach to mitigate dinoflagellate blooms.

Comparing the diel vertical migration of Karlodinium veneficum (dinophyceae) and Chattonella subsalsa (Raphidophyceae): PSII photochemistry, circadian control, and carbon assimilation.

Diel vertical migration (DVM) is thought to provide an adaptive advantage to some phytoplankton, and may help determine the ecological niche of certain harmful algae. Here we separately compared DVM patterns between two species of harmful algae isolated from the Delaware Inland Bays, Karlodinium veneficum and Chattonella subsalsa, in laboratory columns. We interpreted the DVM patterns of each species with Photosystem II (PSII) photochemistry, rates of carbon assimilation, and specific growth rates. Each species migrated differently, wherein K. veneficum migrated closer to the surface each day with high population synchrony, while C. subsalsa migrated near to the surface from the first day of measurements with low population synchrony. Both species appeared to downregulate PSII in high light at the surface, but by different mechanisms. C. subsalsa grew slower than K. veneficum in low light intensities (≈bottom of columns), and exhibited maximal rates of C-assimilation (Pmax) at surface light intensities, suggesting this species may prefer high light, potentially explaining this species' rapid surface migration. Contrastingly, K. veneficum showed declines in carbon assimilation at surface light intensities, and exhibited a smaller reduction in growth at low (bottom) light intensities (compared to C. subsalsa), suggesting that this species' step-wise migration was photoacclimative and determined daily migration depth. DVM was found to be under circadian control in C. subsalsa, but not in K. veneficum. However, there was little evidence for circadian regulation of PSII photochemistry in either species. Migration conformed to each species' physiology, and the results contribute to our understanding each alga's realized environmental niche.