Molecular Insights into Nutrient-Driven Regulation of Trophic Strategies and Life Cycle Shifts in Bloom-Forming Mixotrophic Dinoflagellates

1. Introduction

Molecular machinery, including mechanisms maintaining signal transduction and gene regulation networks, provides a solid though flexible basis for development of cells and organisms within the entire biological life hierarchy, ranging from a variety of intracellular processes to complete life cycles. Among all living creatures, eukaryotic unicellular organisms (protists) are of particular importance for molecular ecologists as they render a unique opportunity to study these mechanisms at the level of a single cell as a whole organism. Knowledge of protistan molecular biology, including life cycle shifts as adaptations to varying physical and chemical features of the environment, is of paramount significance. This information helps resolve such sophisticated issues as chaotic population dynamics (Benincà et al., 2008; Medvinsky et al., 2015), enhancing the evaluation of the balance between external triggers (abiotic characteristics) and internal (cellular, molecular) drivers of cells and organisms development (Telesh et al., 2019; Cagle, Roelke, 2024). Comprehensive understanding of molecular mechanisms of these processes is crucial for the adequate assessments and forecasts of population growth of the ecologically and economically important protists and other influential microorganisms. Of particular interest are toxic and potentially toxic microalgae that cause harmful algal blooms (HABs).

Nowadays, HABs are intensifying and expanding not only at the regional level, e.g., as results of the excess of nutrients due to aquaculture development and increased discharges of agricultural waste waters and fertilizers (Hallegraeff et al., 2021; Zhang et al., 2023), but also worldwide (Glibert, 2020; Gobler, 2020; Hou et al., 2022; Glibert et al., 2025; Li et al., 2026). They occur in a variety of marine and continental water basins of different types, from small eutrophic ponds to vast oligotrophic oceanic regions, since many of those areas are strongly affected by climate change and growing anthropogenic eutrophication (Brown et al., 2020; Anderson et al., 2021; Orlova et al., 2022). The devastating effects of HABs on aquatic ecosystems and their biota are well known and have been repeatedly reviewed elsewhere (Landsberg, 2002; Gobler, 2020; Anderson et al., 2021, 2025; Hou et al., 2022; Dai et al., 2023; Skarlato, Telesh, 2024; Telesh et al., 2025a). In brief, usually by the end of a HAB, the decomposition of algal biomass accumulations results in oxygen depletion in the water column and deterioration of the quality of natural waters, which destabilizes the environment and causes irreparable harm to the flora and fauna, fisheries and fish farming, aquaculture, tourism and public health. The latter issues, along with direct toxic effects of some bloom-forming protists on aquatic organisms and humans, are generally listed among the major negative consequences of HABs (Orlova et al., 2022; Rattner et al., 2022; Occhipinti et al., 2025). However, the bloom onset most often remains enigmatic and unpredictable because of a number of impartial reasons. On the one hand, it depends on numerous, variable and mutually dependent environmental factors. On the other hand, the contribution of molecular data on bloom-forming species to disclosing the triggering mechanisms and driving forces of emergence, development, timing, magnitude and decay of HABs are largely unappreciated. A lack of this knowledge is the major reason for the insufficient effectiveness of many current prognostic models of HABs.

One of the key HAB-forming groups of protists are dinoflagellates (Dinoflagellata). A great part of ca. 2,500 known morphological species of these microalgae cause massive population outbreaks, or blooms, also called “red tides”, in coastal waters of seas and oceans. Moreover, out of the 200 reliably known morphological taxa of various marine phytoplankton whose cells produce toxins, more than 105 are dinoflagellates (Hallegraeff et al., 2021). Their toxins and other secondary metabolites cause up to 500,000 human intoxications per year, with an average mortality rate of approximately 1.5% (Wang, 2008). Besides, among microorganisms, dinoflagellates are the only group of plastid-containing protists capable of bioluminescence: nearly 70 species from more than 18 genera of dinoflagellates express bioluminescence, and the majority of them emit a blue-green light (Okolodkov, 2005, 2011; Marcinko et al., 2013; Timsit et al., 2021).

The effective bloom-forming capacities of many dinoflagellates are backed up by their molecular, cellular, physiological and metabolic peculiarities (Hackett et al., 2004; Okolodkov, 2005, 2011; Soyer-Gobillard, 2025). One of these particular features is the capacity for mixotrophic nutrition and kleptoplasty (Cruz, Cartaxana, 2022), which enhances cells population development often leading to HABs (Millette et al., 2023; Skarlato, Telesh, 2024; Telesh et al., 2025a, 2025b). However, little numerical data on the contribution of these specific characteristics to dinoflagellate HABs formation are available so far. Moreover, mathematical models of these devastating events, based on computational approaches, transcriptomic data, laboratory and theoretical experiments, and the analyses of long-term data series, as well as generalizations of the available knowledge on these topics are scarce (Calbet, 2026). Meanwhile, if available, they could have significantly contributed to the development of comprehensive HAB control strategies for environmental management aimed at effective protection of natural aquatic ecosystems and their resources.

In this review article, molecular features of bloom-forming mixotrophic dinoflagellate cells are considered as the basis for their excess proliferation, including the extreme red-tide events. We discuss the deficiency and redundancy of nutrition substrates in water containing nitrogen (N) and phosphorus (P), and their roles for dinoflagellates population growth and diversity studied in the experiments as well as in nature. We also highlight the effects of variations in nutrient concentrations and availability as modulators of life cycles and nutrition strategies (including mixotrophy and kleptoplasty) of HAB-forming planktonic dinoflagellates. In some sections, the article focuses on biology of a model red-tide microalgae species – the globally distributed mixotrophic, potentially toxic armored dinoflagellates Prorocentrum cordatum (Ostenfeld) J.D. Dodge, 1975, also known by its main synonym Prorocentrum minimum (Pavillard) J.Schiller 1933. Here, we review its peculiar molecular features studied in experiments with laboratory cultures. We also summarize the data on cellular and molecular mechanisms that drive the proliferation of natural P. cordatum populations forming blooms and characterize some of their traits in the southwestern coastal waters of the Baltic Sea.

Special attention is paid to the results of the transcriptomic analyses of molecular machinery involved in life cycle regulation of dinoflagellates, and molecular basics of their mixotrophic strategies in the changing environment. In conclusion, we highlight the dire necessity of including molecular data into conceptual and mathematical ecological models, and particularly the models of kleptoplasty in bloom-forming mixotrophic dinoflagellates, for proper evaluating and forecasting HABs. This approach is in the mainstream of the translational aquatic ecology (TAE) – one of the recently emerged scientific disciplines, which capitalizes on smart conversion of the fundamental ecological findings and hot-spot molecular biology discoveries conducive to their practical use in the essential human life activities: biotechnology, aquaculture, fisheries, environmental management, and nature protection (Skarlato et al., 2018). The authors believe that this review could provide hints to enhancing the linkages between cellular and molecular biology, omics, aquatic ecology and ecological modeling, since merging the approaches of these disciplines is a promising perspective for the future studies that will secure the progress of molecular ecology within the TAE framework.

2. Major Roles of Dinoflagellates in Aquatic Ecosystems

Dinoflagellates form an important marine and brackish-water phytoplankton cluster characterized by pronounced taxonomic and functional diversity. They are crucial to aquatic ecosystems due to high abundances, biomass and productivity. Dinoflagellates effectively fulfill nutrient cycling and energy transfer, synthesize specific toxins, implement benthic-pelagic coupling, form harmful red tides, and thus have huge overall ecological significance (Okolodkov, 2005, 2011; Bravo, Figueroa, 2014; Khanaychenko et al., 2019; Deng et al., 2023; Telesh et al., 2025a, 2026). Numerous studies prove that dinoflagellates play a key role in microplankton biodiversity (Hällfors, 2004; Gómez, 2005; Okolodkov, 2005, 2011). These protists account for more than 2500 actual morphological species and ca. 4000 fossil species (Taylor et al., 2008; Hoppenrath, 2017). The maximum taxonomic diversity of dinoflagellates and their highest abundances are usually registered in marine coastal waters and river mouths (Taylor, Pollingher, 1987; Graham, Wilcox, 2000).

Based primarily on molecular-phylogenetic studies of dinoflagellates, cell biology, modern taxonomy and systematics attribute these protists to a large monophyletic group, Alveolata, along with ciliates (Ciliophora) and sporozoa (Apicomplexa) (Cavalier-Smith, 1991; Fast et al., 2002; Adl et al., 2019). Molecular phylogeny proves that dinoflagellates are most closely related to Apicomplexa but not to Ciliophora (Leander, Keeling, 2003). Meanwhile, the evolutionary line of Alveolata belongs to a supergroup SAR, along with Stramenopila and Rhizaria (Burki et al., 2007). The latter supergroup was rather recently expanded to TSAR due to the addition of the phylum Telonemia after new molecular phylogeny data were obtained (Tikhonenkov et al., 2022).

Most peculiar features of the dinoflagellate cell organization comprise specificity of cellular coverings (Morrill, Loeblich, 1983; Netzel, Dürr, 1984; Pozdnyakov, Skarlato, 2012), unusual structure of the nucleus and permanently condensed chromosomes with distinct chromatin architecture (Rizzo, 2003; Golyshev et al., 2018; Wong et al., 2019). They are also characterized by variability and specific organization of ion channels that shed light on cell physiology (Pozdnyakov et al., 2018, 2020, 2021), and complex structure of chloroplasts (Keeling, 2010; Maréchal, 2024). Molecular features of dinoflagellates involve one of the largest genomes among the eukaryotes, extensive gene duplication, peculiar gene organization patterns such as tandem arrays, specific features of splicing, predominance of post-transcriptional and translational regulation over transcriptional control, and other processes (Bachvaroff, Place, 2008; Lin, 2011, 2024; Fukuda, Suzaki, 2015; Berdieva et al., 2026). These microalgae have a complex life cycle with the alternating vegetative and sexual phases that employ various strategies and include dormant immotile stages (Bravo, Figueroa, 2014; Figueroa et al., 2018). The sexual process in the life cycle of dinoflagellates is known and generally regarded as both a survival strategy in the adverse environmental conditions like nutrient deficiency, stressful temperature and salinity regimes, intense grazing, or parasite infection (Agrawal, 2012; Kalinina et al., 2023, 2025), and a proliferation strategy (Lin et al., 2025).

Among functional traits, the ability of dinoflagellates to symbiotic relations with corals is well known (Aranda et al., 2016). In marine and coastal ecosystems, dinoflagellates are crucial being major primary producers along with diatoms (Fensome, Munsterman, 2026). Dinoflagellates are an effective biological “carbon pump” and a means of transporting nutrients within the food web, serving as food for microzooplankton and fish larvae (Winder, Sommer, 2012; Mitra et al., 2014; Khanaychenko et al., 2019). They can form devastating HABs in marine and brackish-water coastal areas worldwide (Heil et al., 2005; Orlova et al., 2014, 2022; Li et al., 2022), including the Arctic seas (Okolodkov, 2005; Silva et al., 2023). In addition, it has been established that the biomass of these protists during the blooming period can be used for thermochemical conversion production of liquid biofuels with its subsequent catalytic purification (Zhou, Hu, 2020). On a global time scale, because of the mass decomposition of algal cells after the cessation of the bloom, thecae of dinoflagellates accumulate in bottom sediments and contribute to the formation of fossil fuels (Okolodkov, Cervantes-Urieta, 2026).

Many dinoflagellates produce structurally diverse toxins that are derived via divergent pathways (Tango et al., 2005; Grattan et al., 2016; Verma et al., 2019; Guillotin, Delcourt, 2022). The main toxin-producing dinoflagellate species in plankton belong to the genera Prorocentrum, Alexandrium, Dinophysis, Gambierdiscus, Ostreopsis and Karenia. The dinoflagellate toxins cause nausea, vomiting, abdominal pain, diarrhea, and neurologic symptoms such as fatigue, myalgias, pruritus, circumoral paresthesias, reversal of hot and cold sensation, psychiatric abnormalities, and memory loss; the neurologic symptoms may persist for months to years (Wang, 2008).

Along with toxins sensu lata, marine dinoflagellates also produce a wide range of other biologically active secondary metabolites, including unique pigments like peridinin (Takaichi, 2011) and specific proteins, although the roles of some of them remain poorly understood so far (Pechkovskaya et al., 2021, 2023; Filatova et al., 2025). As demonstrated for various microalgae, some of these compounds possess antiviral (Carbone et al., 2021), antibacterial (Najdenski et al., 2013), antifungal (Li et al., 2006), and antitumor (Martínez Andrade et al., 2018) properties; therefore, they can be used in pharmacology and food industries (Pulz, Gross, 2004; Borowitzka, 2013). Much attention has been paid to dinoflagellates’ metabolites containing complex compounds with diverse chemical natures (Assunção et al., 2017), including macrolides, cyclic polyethers, spirolides, purine alkaloids, and other components (Kellmann et al., 2010).

Currently, dinoflagellates are being actively used in the search for new natural migrastatic substances that could inhibit cancer cells motility, thereby preventing metastasis. For example, recent studies using the MALDI-TOF technique (Calvano et al., 2015) have identified biologically active secondary metabolites, including pigments chlorophyll a and peridinin, and proteins (chaperones, cytoprotective and antioxidant proteins), produced by the potentially toxic dinoflagellates Prorocentrum cordatum (Filatova et al., 2024, 2025). Specifically, using adherent murine hepatoma MH22a cells as one of the target models it was found that the ethanol extract of P. cordatum, due to its low toxicity and ability to suppress cell adhesion and migration, appeared to be the most promising candidate for further studies as a potential biologically active agent (Filatova et al., 2025).

In aquatic ecosystems, dinoflagellates also facilitate connectivity between the pelagic and benthic communities by producing cysts that accumulate on the bottom but re-inoculate the water column when favorable conditions for excystation are restored (Bravo, Figueroa, 2014; Skarlato et al., 2018; Liu et al., 2020). The life cycle complexity of these protists defines the variability of their dormancy strategies; therefore, duration of the immotile persistence of dinoflagellates in the ecosystem is determined by the type of cysts. The thick-wall long-term dinosporin-containing resting cysts are characterized by a significant period of dormancy, which can reach several months or even years (Bravo, Figueroa, 2014). The thin-wall — pellicle or thecate — temporary cysts do not undergo prolonged dormancy and stay immotile for a few hours or days, although sometimes for months (Bravo et al., 2010; Bravo, Figueroa, 2014; Matantseva et al., 2020). Consequently, the nature of the benthic-pelagic coupling involving HAB-forming dinoflagellates is complex and ambiguous, and these interactions play a dual role in the formation of red tides: first, due to cyst formation and deposition at the bottom and, second, because of the composite, non-linear effects of benthic feeding on nutrients content in water. Concerning the latter, on the one hand, the excretion of available nutrients by organisms in benthos such as bivalves, accompanied by the excystation of dinoflagellates under favorable conditions and their release into plankton, increases the trophic status of a water body and promotes HABs. On the other hand, the elimination of microalgae by these same mollusks during their intense feeding by filtration reduces the population size of bloom-forming species and thus prevents red tides (Telesh et al., 2025a).

Many HAB-forming planktonic dinoflagellates thrive in various aquatic environments and withstand external stressors due to their wide ecological niches, like the cosmopolitan marine mixotrophic species Prorocentrum cordatum, particularly with respect to temperature, pH, salinity, and concentration of basic nutrients (Telesh et al., 2016). Besides, largely due to mixotrophy of many dinoflagellate species (Stoecker et al., 1991, 1997, 2017; Matantseva, Skarlato, 2013), the broad nutrition spectrum is another great ecological advantage of these protists (Jeong et al., 2010, 2021). Mixotrophic dinoflagellates utilize a wide range of organic and inorganic resources available for their growth and reproduction, often changing trophic preferences depending on environmental conditions (Abassi, Ki, 2022; Li et al., 2026). Specifically, by using stable isotope tracers and NanoSIMS, it was shown that individual P. cordatum cells within a population displayed significant heterogeneity in the rates of nutrient uptake and extent of the urea-mediated inhibition (by 30-40%) of the nitrate uptake (Matantseva et al., 2016). Thus, starting to consume urea at a high rate as soon as it appeared in the environment, P. cordatum demonstrated the rates of urea-N uptake that were 1.6-4.1 times higher than those of nitrate-N in P. cordatum population (Matantseva et al., 2016). Moreover, these dinoflagellates also demonstrated the uncoupled assimilation of carbon and nitrogen from urea and the amino acid glycine (Matantseva et al., 2018). The discovered functional heterogeneity of these protist populations, leading to the putative co-existence of various sub-populations with differing nutrition modes, ensures the ecological success of these dinoflagellates and enhances their competitive advantages in stressful environments.

The above features contribute to the proliferation of dinoflagellates in diverse habitats, in water bodies of different types, and the expansion of red tide zones therein (Figure 1). They also allow these protists to compete successfully with closely related species, as demonstrated by the invasion history of P. cordatum in the Baltic Sea and its displacement of the previously dominant congeners among planktonic dinoflagellates therein (Telesh et al., 2016). Thus, knowledge of the nutrition strategies of these flagellates and new data on their ecological niches have already made a significant contribution to HAB research (Telesh, Skarlato, 2022). Specifically, it was shown that different trophic strategies of these microorganisms determine their ability to rapidly proliferate by modulating growth rate and reproductive efficiency, providing competitive advantages, high invasion potential and range expansion (Matantseva et al., 2016, 2018; Telesh et al., 2016, 2021, 2024, 2025a, 2025b, 2026).

Enumeration of peculiar molecular, cellular and physiological features and functional traits of dinoflagellates can be continued, and their list can be much longer. However, we will cease it for now, since the subsequent sections of this review will focus primarily on the essential nutrients and their roles in the molecular machinery of dinoflagellate trophic strategies, reproduction and life cycle regulation. All together, these characteristics secure pronounced adaptability of these protists to climate changes and anthropogenic stresses; enhance their persistence, population growth, and HABs formation.

3. Nitrogen and Phosphorus as Essential Nutrients Modulating Dinoflagellate Cell Physiology

Nitrogen (N) and phosphorus (P) are the two biogenic elements whose availability in the environment constitutes one of the principal prerequisites regulating various aspects of dinoflagellate cell physiology. Nitrogen is a structural component of amino acids, nucleotides, and chlorophyll molecules. The availability of nitrogen limits the productivity of marine microbial communities and plays an essential role in structuring of phytoplankton species composition (Bertrand, Allen, 2012; Moore et al., 2013; Shibata et al., 2015). Marine phytoplankton provide a major proportion of global primary production and modulate biogeochemical cycles by assimilating nitrogen. It is usually assumed that most microorganisms can use dissolved inorganic nitrogen (DIN) in the form of nitrate, nitrite, and ammonium (Zehr, Ward, 2002). Dissolved organic nitrogen (DON) can be a large pool in the oceans and an even larger one in coastal waters. A significant fraction of this pool is composed of amides (including urea); it also includes heterocyclic compounds, proteins and amino acids (Zehr, Ward, 2002). Nitrogen limitation affects metabolic processes in dinoflagellate cells, enhancing glycolysis, the pentose-phosphate pathway, the tricarboxylic acid cycle, and fatty acid metabolism, and altering expression of genes encoding photosystem proteins (Cooper et al., 2016; Hou et al., 2018; H. Li et al., 2021; T. Li et al., 2021; Shi et al., 2021). Nitrogen deficiency activates nitrate/nitrite and urea transporters to increase assimilation of available nitrogen sources (H. Li et al., 2021; Shi et al., 2021).

Phosphorus is a biogenic element, which plays a crucial role in the cells as it is a component of nucleic acids (DNA, RNA), cell membranes (phospholipids), signaling molecules (inositol triphosphate, cAMP), nicotinamide adenine dinucleotide (NAD) and its phosphorylated form (NADP), adenosine triphosphate (ATP, an energy currency), etc. Phosphorylation activates many enzymes necessary for maintaining basic metabolic pathways and regulating the cell cycle. Thus, phosphorus availability has a significant impact on the viability and growth of marine phytoplankton and, consequently, on their primary production in the seas and oceans.

Phosphorus can be available to dinoflagellates in the form of dissolved inorganic phosphate, DIP (primarily orthophosphate [Pi]) and as a variety of dissolved organophosphorus (DOP) compounds such as ATP, cytidine-5-monophosphate, fructose-6-phosphate, glucose-6-phosphate, glycerophosphate, uridine-5-monophosphate, phenyl phosphate, RNA, etc. (Antia et al., 1990; Huang et al., 2005; Oh et al., 2010; Li et al., 2015; Lin et al., 2016). DOP appears to be the main source of this nutrient during algal blooms, when inorganic nutrients are depleted (Huang et al., 2022). Phosphorus deficiency stimulates phosphate uptake and intracellular recycling (Lei, Lu, 2011; Morey et al., 2011; Zhang et al., 2014; Li et al., 2016). For example, proteomic analysis of phosphate starved Karenia mikimotoi revealed overexpression of vacuolar phosphate transporters, the proteins associated with vesicular transport and proteins responsible for lipid and phospholipid degradation (Lei, Lu, 2011). K. mikimotoi and Prorocentrum shikokuense Y.Hada[1] have also been shown to replace membrane phospholipids partially with glycerol glycolipids and betaine lipids (Huang et al., 2023; Li et al., 2024). In freshwater ecosystems, despite the historical “dogma” defining them as strictly phosphorus-limited (Hecky, Kilham, 1988), many systems exhibit co-limitation, where both N and P availability simultaneously constrain primary production (Bracken et al., 2015; Lewis et al., 2020).

Notably, toxin biosynthesis depends on availability of both elements. Primarily, nitrogen is required for the production of saxitoxins, as shown for Alexandrium tamarense and Alexandrium minutum (Béchemin et al., 1999; Chen et al., 2024). The most recent studies indicated that high nitrogen supply enhanced not only saxitoxin biosynthesis, but also toxin release: excess nitrogen promoted sxtA4 gene expression by Alexandrium catenella (Whedon & Kofoid) Balech, with a positive correlation between sxtA4 and total yield (Lian et al., 2026). Nevertheless, the production of okadaic acid (diarrheic shellfish toxin) is modulated by phosphorus level (Chen et al., 2025).

4. Trophic Strategies of Dinoflagellates in the Changing Environment: Mixotrophy and Kleptoplasty as Its Peculiar Constituent

In recent years, researchers have been paying much attention to studying not only the ecological prerequisites of dinoflagellate blooms but also the intracellular drivers and molecular mechanisms that back up their trophic strategies and secure fast population growth often leading to HABs (Soyer-Gobillard, 2025). Mixotrophy is one of the nutrition modes typical for many bloom-forming dinoflagellates that combine autotrophy due to photosynthesis in the light using inorganic nutrient substances with heterotrophy in the forms of osmotrophy and phagotrophy (Stoecker, 1991; Stoecker et al., 1997, 2017; Matantseva, Skarlato, 2013; Mansour, Anestis, 2021). Osmotrophy involves the consumption of dissolved organic compounds by osmosis, and phagotrophy is the ability of cells to engulf, absorb and assimilate solid organic substances, such as microalgae and bacterial cells. Currently, photo-osmo-mixotrophy is considered a generally accepted and widespread nutritional mode of microalgae (Mitra et al., 2023). At the same time, mixotrophy, which includes phagotrophy, is not ubiquitous; moreover, the modes of use of this nutritional strategy also vary significantly among diverse planktonic protists (Mitra et al., 2016, 2023).

The combination of the two strategies, photo-osmo-mixotrophy and photo-phago-mixotrophy, provides high adaptability of protists to fluctuations in the composition and abundance of food resources and, consequently, increases the ecological homogeneity of many unicellular mixotrophic organisms, which provides the basis for their unification within the concept of “mixoplankton” (Flynn et al., 2013, 2019). Mixoplankton (i.e., plankton capable of mixotrophy) are generally characterized by high adaptability to fluctuating environment (Glibert, 2015; Wilken et al., 2020). This is primarily due to the ability of mixoplankton to utilize various alternative resources (Matantseva et al., 2016, 2018), and their broad ecological niches, including a wide range of food substrates (Telesh et al., 2016, 2024; Jeong et al., 2021). As a result, because of the high adaptability and rapid growth of mixoplankton populations, they are often the reason of HABs (Anderson et al., 2019; Carstensen et al., 2020; Glibert, 2020; Hallegraeff et al., 2021; Karlson et al., 2021).

Currently, an increasing number of dinoflagellate species are exhibiting mixotrophic properties, and this enhances their resilience to changing environmental conditions, including shifts in the composition and concentration of nitrogen- and phosphorus-containing compounds or decreased light, which impedes photosynthesis. Specifically, phosphorus deficiency stimulates phagotrophy in some mixotrophic species of dinoflagellates, such as Prorocentrum cordatum and Tripos sp. (the latter was described as Ceratium furca (Ehrenberg) Claparède & Lachmann) (Smalley et al., 2003; Johnson, 2015; Berdieva et al., 2020), and increases expression and activity of cell-surface alkaline phosphatase (Dyhrman, Palenik, 1997; Lin et al., 2012; Ou et al., 2020). A lack of phosphorus in the environment induces the expression of plasma membrane and vacuolar phosphate transporters as well as proteins associated with vesicular transport (Lei, Lu, 2011; Kalinina et al., 2025). Nitrogen depletion can also induce phagotrophy in P. cordatum (Johnson, 2015) and Karlodinium veneficum (D.Ballantine) J.Larsen (described as Gyrodinium galatheanum (Baarud) Taylor in: Li et al., 2000), although to a lesser extent than phosphorus. Activation of endocytosis and phagosome-related pathways has also been detected at the transcriptomic level (H. Li et al., 2021).

Recently, it has been shown that none of the dominant HAB-forming dinoflagellate species in the coastal waters of the southwestern Baltic Sea is purely autotrophic since seven out of ten of these dominants are mixotrophs, while the remaining three species are heterotrophs (Telesh et al., 2026), following the classification by Jeong with co-authors (2021). Mixotrophy involving phagotrophy is observed in all phytoplankton clades, with the exception of diatoms and cyanobacteria (Borics et al., 2021). Moreover, nowadays mixotrophy is considered not just a nutritional strategy, but also a unique life strategy for dinoflagellates and many other protists (Jeong et al., 2021; Chen et al., 2025; Glibert et al., 2025). The environmental triggering of mixotrophy is highly complex and multidimensional (reviewed by Calbet, 2026). Recent transcriptomic analyses have revealed that mixotrophy is not just the synergistic process of “grazing while photosynthesizing”; rather, prey availability can trigger large-scale transcriptional reprogramming that shifts cellular metabolism toward heterotrophic pathways (Lie et al., 2017; Ma et al., 2023; Calbet, 2026). However, some subtle molecular and cellular aspects of mixotrophic metabolism remain poorly understood (Glibert, Mitra, 2022).

Kleptoplasty is one of the less studied though most enigmatic intracellular mechanisms of mixotrophy (Nishitani et al., 2011). The term “kleptoplasty” refers to the processes of acquisition, accumulation, and utilization of kleptoplasts, i.e., unassimilated chloroplasts of microalgae absorbed by heterotroph and mixotroph cells during phagotrophic nutrition (Gast et al., 2007; Rumpho et al., 2007; Kim et al., 2012; Millette et al., 2023). Kleptoplasty is observed in a wide range of not only unicellular organisms such as dinoflagellates, ciliates, and foraminifera (Stoecker et al., 2009; Pillet, Pawlowski, 2013), but also in multicellular invertebrates, for example, sacoglossan sea slugs of the genus Elysia (Rumpho et al., 2007), and some flatworms (Van Steenkiste et al., 2019).

It has been established that kleptoplasts not just persist, but also photosynthesize in the host cell, sometimes for a relatively long period, up to several months (Stamatakis et al., 2017), after which the photosynthesis of kleptoplasts is diminished and finally lost, and/or the “stolen” plastid is digested (Mansour, Anestis, 2021). For example, in Nusuttodinium aeruginosum (F.Stein) Y.Takano & T.Horiguchi, kleptoplasts can be inherited by the daughter cells through five generations, and can even be found in the resting cysts (Onuma, Horiguchi, 2015). Along with their own plastids, kleptoplasts in mixotrophs serve as additional “reactors of photosynthesis” (Skarlato, Telesh, 2024; Telesh et al., 2025b). For the majority of photophago-mixotrophic protists, however, the question of how great the contribution of functioning kleptoplasts to the overall photosynthesis and cell growth is – remains open.

Dinoflagellates obtain kleptoplasts from prey algae such as various representatives of cryptophytes (Li et al., 2000), haptophytes (Berge et al., 2008), chlorophytes (Strom, Buskey, 1993), prasinophytes (Berge et al., 2008), raphidophytes (Nakamura et al., 1995), diatoms (Menden-Deuer et al., 2005; Yoo et al., 2009), and other dinoflagellates (Tillmann, 2004; Adolf et al., 2006). The pathway of plastid acquisition can be intricate. Dinophysis acuminata Claparède & Lachmann acquires plastids of Geminigera/Teleaulax cryptophyte from the captured ciliate Myrionecta rubra (Lohmann) Jankowski (Wisecaver, Hackett, 2010). As a result, the two outer plastid membranes, the cryptophyte nucleus and the nucleomorph, are lost. At the same time, the nuclear genome of D. acuminata contains five plastid genes from multiple algal lineages, only one of which was derived from a cryptophyte, and this implies instability of the photosynthetic system in these dinoflagellates (Wisecaver, Hackett, 2010). Meanwhile, the Antarctic Ross Sea dinoflagellate appears to retain its ancestral secondary plastid and obtains temporary kleptoplasts stolen from haptophyte prey partitioning functions between the two plastid’ systems (Hehenberger et al., 2019). Moreover, this dinoflagellate has acquired nuclear genes by horizontal gene transfer (HGT) from outside the kleptoplast lineage; however, those HGT-genes encode kleptoplast-targeted proteins. Their expression is not strongly changed by environmental conditions and many of them are shared with related dinoflagellates with fully integrated plastids. Such a pattern suggests that the protein targeting system had developed before the plastid became permanently fixed (Hehenberger et al., 2019).

In this regard, a separate issue is the fate and physiological role of captured plastids in a mixotrophic dinoflagellate cell. Laboratory experiments showed that, for example, for Prorocentrum spp. (P. micans Ehrenberg, P. cordatum), the cryptophyte microalgae are most often the optimum prey in terms of cell size and consumption rate (Jeong et al., 2010). The results of the long-term Baltic Sea database analyses support this outcome by demonstrating the co-existence of P. cordatum and P. micans with cryptophyte species like Hemiselmis virescens Droop or Teleaulax sp. in plankton, resulting in strong and statistically significant (p < 0.05) positive correlations between the abundance of Prorocentrum spp. and the cryptophytes (Table 1).

In the meantime, neither the laboratory studies of the dinoflagellates feeding rates nor the field data on their co-occurrence with certain prey objects can provide information on the efficiency of kleptoplasty, while the experimental investigations of this specific and delicate intracellular process are problematic yet. Thus, in most cases, the real contribution of kleptoplasty to dinoflagellate cells growth remains unknown; therefore, this phenomenon has not been taken into account in the majority of available mathematical models of the growth and population dynamics of mixotrophic organisms (Flynn, Mitra, 2009; Mitra et al., 2014; Chen et al., 2025). The only exception known so far is a mathematical model of kleptoplasty based on the fundamental J. Monod model of microorganisms growth (Monod, 1949), a series of theoretical experiments (“thought experiments”), and a set of differential equations for a simple biological system, which consisted of one mixotrophic protist cell, one inorganic resource, and one organic resource (Telesh et al., 2025b). The model was developed and tested using experimental data on the feeding of dinoflagellates Prorocentrum cordatum on the cryptophyte microalgae Teleaulax sp. (Jeong et al., 2010, 2021), taking into account their reproduction rates and morphometry (Heil et al., 2005). This study also allowed the authors to suggest an original kleptoplasty index (k) – a new ecological indicator reflecting the quantitative contribution of kleptoplasty as an important intracellular process to HABs formation. In particular, the model has shown that kleptoplasty of the mixotrophic P. cordatum using Teleaulax sp. as food can increase the cell division rate by up to 40%, promoting rapid population growth of these dinoflagellates and the subsequent algal bloom (Telesh et al., 2025b).

In the future modeling studies, the assessment of the role of kleptoplasty in the formation of dinoflagellate HABs can be correlated with the major cellular parameters and molecular mechanisms that determine the functioning of these protists. Thus, further research into molecular basics of kleptoplasty as one of the key mixotrophic nutrition mechanisms affecting dinoflagellate HABs can be a future prospect of molecular ecology, which could help understand algal blooms and contribute to prognostic mathematical modeling of HABs.

5. Effects of Nitrogen Redundancy on the Expression of N-Transporters’ Genes, Transcriptional Responses, Diversity, and Dinoflagellates Population Growth

The abundance, composition and proportions of different inorganic and organic nutrients as food substrates in the environment play a crucial role in mixoplankton development and proliferation. The presence of different forms of nitrogen is mandatory for various cellular metabolic processes of dinoflagellate cells that secure their maintenance, growth, and reproduction (see also Section 6). For example, experimental results suggest that mixotrophic dinoflagellates Prorocentrum cordatum are sensitive to the proportions of N-containing substrates and can adopt the most effective strategy for assimilation of available nitrogen sources in the changing environment, demonstrating response to shifts in their composition (Matantseva et al., 2016, 2018; Berdieva et al., 2021a; Pechkovskaya et al., 2020; Kalinina et al., 2023). It was discovered that the expression levels of the genes dur3 and nrt2 encoding the transporters of urea and nitrate, respectively, in the NO₃-grown cultures depended on the added nitrogen sources being the lowest in the presence of ammonium (Pechkovskaya et al., 2020).

Studies of the transcriptional responses of P. cordatum to available N-containing substrates allowed estimating the influence of NO₃⁻, NH₄⁺ and urea on the transcription levels, as well as on the expression of urea and nitrate transporter genes in these dinoflagellates. Specifically, it was shown that dur3 expression levels were downregulated after the supplementation of additional N sources and were 1.7–2.6-fold lower than in the nitrate-grown culture, while the nrt2 expression levels decreased 1.9-fold in the presence of NH₄⁺ (Pechkovskaya et al., 2020). Total RNA and DNA synthesis rates, estimated by the analysis of incorporation of ³H-thymidine and ³H-uridine in batch and continuous cultures, have demonstrated that the addition of N compounds did not affect the DNA synthesis rates, while the transcription levels increased up to 12.5-fold after the excessive N supplementation (Pechkovskaya et al., 2020). These investigations of various nitrogen sources as biomarkers of dinoflagellate proliferation due to their differentiated impact on expression of dur3 and nrt2 genes and modification of transcription rates in the mixotrophic P. cordatum allowed concluding that the knowledge on nitrogen redundancy, diversity and proportions of N-substrates in the environment is of paramount importance. Incorporation of these data into ecological models could increase the precision of the future HAB forecasts under global N pollution and eutrophication.

Meanwhile, along with different nutrient sources, natural systems are characterized by fluctuating light, episodic nutrient pulses, variable prey communities, and shifting thermal gradients; therefore, the kinetics of trophic switching in mixoplankton under such dynamic conditions and their effects on cells proliferation remain poorly characterized (Calbet, 2026). Besides, inter- and intra-population heterogeneity of cells as well as the ambiguity of dinoflagellates’ behavior under various stresses (Matantseva et al., 2020; Pechkovskaya et al., 2021) and their peculiar life cycles (Kalinina et al., 2018, 2020, 2023; Berdieva et al., 2020, 2021a,b) hamper the usage of the available data on dinoflagellate cell biology for prognostic mathematical modeling of their HABs. Furthermore, the molecular mechanisms linking environmental sensing to the coordinated regulation of phototrophic and heterotrophic machinery are only just starting to be mapped (Calbet, 2026).

Nevertheless, certain advances in unveiling the linkage of some environmental prerequisites and fine cellular-molecular mechanisms underlying dinoflagellate blooms have been achieved in recent decades. For example, it has been shown that the concentration of total nitrogen (TN) in water affects development and growth of protist cells as well as the overall phytoplankton composition, abundance and distribution; thus, TN can be considered an anthropogenic stress marker (Rönnberg, Bonsdorf, 2004; Howarth, 2008; Schiewer, 2008; Mekonnen, Hoekstra, 2015; Zhang et al., 2023). Recently, it has also been demonstrated that dinoflagellates with the widest TN-niche limits (along with broad niches for total phosphorus, water temperature and salinity) have the strongest potential for most frequent bloom events in the fluctuating coastal environments (Telesh et al., 2024). The latter study suggested a conceptual model for merging the bloom-forming capacity of dinoflagellates with the width of their TN niches (Figure 1), and this approach allowed coming closer to prediction of red tides.

Broad range of organic and inorganic resources used by mixoplankton underpins coexistence of quite a large number of mixotrophic dinoflagellates in space and time, as demonstrated by the data from the Baltic coastal waters (Telesh et al., 2016, 2021, 2024, 2025b). Meanwhile, our recent research hypothesis, suggesting those coastal regions with TN concentrations that are optimal for bloom-forming dinoflagellates to host greater taxonomic diversity of these protists, compared to the areas with non-optimum TN content, unexpectedly was not supported by the analyses of the 82 dinoflagellate species in the 44 years-long database from the SW Baltic Sea (Telesh et al., 2026). Instead, surprisingly, the highest species richness of these planktonic protists was observed at relatively low eutrophication level (TN < 15 µmol/L). Moreover, the study has demonstrated that the diversity of dominant bloom-forming dinoflagellate species decreased gradually from low to moderate, high, and very high eutrophication levels, with the minimum number of species in the extremely eutrophic waters at TN > 45 µmol/L (Telesh et al., 2026). These results disagree with the theoretically expected species-richness maximum at the intermediate TN concentrations, as predicted by the intermediate disturbance hypothesis (IDH), which postulates that maximum species richness should be expected at moderately stressed conditions (Connell, 1978). A possible though speculative explanation of this discrepancy could involve the linkage between the dinoflagellate growth rates and TN disturbance frequencies rather than magnitude (Gaedecke, Sommer, 1986; Weider, 1992). As shown earlier, usually a species diversity maximum is observed when disturbance frequencies are equal to a three-generation cycle (Lampert, Sommer, 2007). For dinoflagellates at nutrient-saturated (i.e., optimum) conditions, the growth rates between ca. 0.3–0.7 day^-1 are known from laboratory experiments (Jia et al., 2019), which is in accord with field observations (Baek et al., 2008) and the knowledge that at suboptimal conditions, the algal growth rates decrease. Consequently, this will shift the optimum disturbance pattern from ca. 4–10 days to much longer periods, allowing for other disturbance factors than TN to serve as stressors affecting the dinoflagellates competition pattern. In turn, slower growth of the superior competitor generally favors the enhanced survival time of the inferior ones. In such a case, the relation between these parameters could secure different competition patterns at the optimal and suboptimal TN conditions, promoting higher species richness of mixotrophic dinoflagellates at suboptimal TN concentrations due to lower growth rates and, therefore, less effective competition for nutrients (Telesh et al., 2026).

Thus, similarly to the ambiguous effects of benthic-pelagic coupling involving HAB-forming mixotrophic dinoflagellates (Telesh et al., 2025a), the effects of nitrogen redundancy on dinoflagellate diversity is also nonlinear and enigmatic. This fact demonstrates that nutrient-driven regulation of biodiversity and trophic strategies, including mixotrophy, is a dynamic, multi-faceted stoichiometric optimization, which is shaped by cells assimilation efficiency, functional architecture of aquatic assemblages, and the complex ever-changing reality of the marine environment (Calbet, 2026). Therefore, the role of TN excess and overall nitrogen redundancy in dinoflagellate cells growth, gene expression, transcriptional responses, and population development of these mixoplankters requires further investigations. Prospective studies could include testing the applicability of Tilman’s resource competition theory (RCT; Tilman, 1977, 1980; Zhang, Becks, 2025), aiming to unveil the unexpected response of dinoflagellate biomass to N-availability, as described above. Previously, the applicability of RCT has been shown by both theoretical and practical approaches (e.g., Smayda, Reynolds, 2003; Bi et al., 2021), at least for the competitive relations between dinoflagellates and diatoms. However, with respect to nitrogen as a co-limiting resource, the competition of diatoms and dinoflagellates with cyanobacteria still needs to be unravelled. The existing knowledge refers to several potential mechanisms ranging from vertical migration (e.g., Fisher et al., 2013) to allelopathic interactions (e.g., Vardi et al., 2002). However, a comprehensive study, which also takes into account the variable and complex species-specific trophic strategies (Smayda, Reynolds, 2003), is still lacking.

6. Nitrogen and Phosphorus Deficiency as Major Stressors Modulating Life Cycles of Dinoflagellates

Nitrogen and phosphorus deficiency limits cell proliferation and can lead to cell cycle arrest, as shown for various dinoflagellate species (e.g., Blanco, 1995; Zhang et al., 2014; Li et al., 2016; Zhao et al., 2017; Chen et al., 2024). Particularly, phosphorus deficiency leads to cell cycle arrest in G1/G0 phase, although the cells continue to grow and increase in size (Lei, Lu, 2011; Zhang et al., 2014; Li et al., 2016). At the molecular level, the lack of nutrients suppresses cell-cycle regulators such as G1/S-specific cyclin and cyclin-dependent kinases (Shi et al., 2017; H. Li et al., 2021). In Alexandrium catenella, phosphorus deficiency reduces expression of calcium-dependent protein kinase (CPK), which plays a crucial role in signal transduction and cell division in plants. In contrast, fizzy/cell division cycle 20-related protein (FDC) is upregulated; it acts as a negative regulator of the cell cycle, inhibiting cell division by regulating the anaphase-promoting complex (APC) and maintaining the required levels of mitotic proteins (Zhang et al., 2014). More importantly, nitrogen and phosphorus depletion serves as a notable signal that initiates life-cycle transitions promoting long-term population survival.

Phosphorus deficiency can trigger the transfer to sexual phase in complex life cycles of diverse marine dinoflagellates (Supplementary Materials, Table S1). Among those species are Prorocentrum cordatum, Lingulaulax polyedra (syn. Lingulodinium polyedra), Gymnodinium nolleri, Protoceratium reticulatum, and Alexandrium minutum (Figueroa, Bravo, 2005; Figueroa et al., 2006c; Figueroa et al., 2011; Salgado et al., 2017; Kalinina et al., 2023). These processes included gamete formation and fusion, planozygote formation and encystment.

Life cycle transition has been well described in the clonal culture of P. cordatum, where two weeks of phosphate deprivation led to increase in the proportion of planozygotes with relative nuclear DNA content 2C and 4C (Kalinina et al., 2023). Transcriptomic analysis at this stage revealed increase in the expression of six homologs of meiosis-associated genes, that are rad51, spo11, RAD21/REC8-like protein coding gene, exo1, msh4 и msh5, responsible for meiotic recombination and double-strands break DNA reparation (Kalinina et al., 2025). The addition of full medium containing sufficient amounts of all the nutrients stimulated cell divisions and transition to vegetative 1C phase (Kalinina et al., 2023).

Nitrogen limitation also serves as a nutrient-related trigger of transition to the sexual phase (Table S1). In laboratory experiments, cultivation under nitrogen deficiency induced sexuality in various marine dinoflagellates — Alexandrium taylori, Karenia brevis, Kryptoperidinium triquetrum (syn. K. foliaceum), Levanderina fissa (syn. Gyrodinium uncatenum), and Scrippsiella lachrymosa (Anderson et al., 1985; Figueroa et al., 2006a; Figueroa et al., 2009; Persson et al., 2013; Persson et al., 2021). In Lingulaulax polyedra, planozygotes encysted in N-depleted medium, as in P-depleted conditions, although nitrogen deficiency yielded more temporary cysts while phosphorus deficiency produced a greater yield of resting cysts (Figueroa, Bravo, 2005). Gymnodinium catenatum showed a similar switch in response to nitrogen deficiency (as well as phosphorus deficiency), but only this condition, like replete medium, allowed most planozygotes to undergo division, and more cysts to successfully germinate and retain viability, despite frequent formation of faulty cyst walls (Figueroa et al., 2006b).

Notably, available literature on freshwater species mostly reports the effects of nitrogen deficiency on reproductive strategy that are consistent with some of the more complex ideas about nutrient regulation in freshwater ecosystems. Several studies describe Peridinium species induced to enter the sexual process, including encystment of planozygotes (Pfiester, 1975, 1977, 1984; Sako et al., 1984; Chapman, Pfester, 1995) (Table S1).

Transcriptomic data reflecting reproductive strategy transitions in response to nitrogen limitations in dinoflagellates also remain sparsely discussed. Nevertheless, Alexandrium minutum has been shown to upregulate some genes coding for meiosis-associated proteins under nitrogen deficiency, for example, homologs of the double-strand break repair protein MRE11, topoisomerase II, DNA mismatch repair protein MLH3, and about two dozen homologs of the RNA-binding protein MEI2 (Akbar et al., 2023). MEI2-like proteins are abundant in dinoflagellates and appear to form two putative functional sets: one related to the sexual phase and another associated with encystment of vegetative cells (reviewed by Berdieva et al., 2026). Two mei2-like genes also altered their expression in nitrogen-starved cells of Amphidinium carterae Hulburt: one was upregulated and the other slightly downregulated (Lauritano et al., 2017).

Nutrient deficiency can specifically trigger the direct transition from vegetative growth to resting cyst formation since cysts can be formed not only as a dormant stage in zygote development, but also as survival strategy when conditions for the vegetative phase are unfavorable. Resting cysts are protected against viruses, predators, and parasites. Nitrogen limitation effect has been shown in some marine armored and naked dinoflagellates, including Alexandrium minutum (syn. A. lusitanicum), A. taylori, Gymnodinium corollarium, Lingulaulax polyedra, and Scrippsiella acuminata (syn. S. trochoidea) (Blanco, 1995; Figueroa et al., 2006a; Kremp et al., 2009; Yue et al., 2022). A slight effect was reported for the peridinialean Ensiculifera sp. (Blanco, 1995). Karenia mikimotoi forms cysts referred to as temporary; however, the presented micrographs do not allow confident identification of their type (Zhao et al., 2017). In freshwater species, nutrient depletion can also stimulate encystment (Chapman, Pfester, 1995). In P-depleted conditions, encystment has been shown in Alexandrium taylori, Lingulaulax polyedra and Gymnodinium catenatum. In the two latter species, phosphorus depletion has a stronger effect on cyst production than nitrogen limitation does (Blanco, 1995; Figueroa et al., 2006a, b), while in Scrippsiella acuminata and Alexandrium minutum, the opposite was true (Blanco, 1995; Yue et al., 2022). Thus, the nutrient deficiency effects cannot be generalized across taxa. Yue et al. (2022) also showed that the life-cycle transition was consistently accompanied by altered expression of cell-cycle regulators, including cyclin B and cyclin-dependent kinase 1; expression of these genes in resting cysts remained moderate but significantly lower than that in fast-growing vegetative cells.

It is noteworthy that cysts must possess sufficient ATP reserves to germinate. For example, S. acuminata cysts can take up phosphate from the environment (Rengefors et al., 1996). This enables dinoflagellate cysts to accumulate phosphorus from benthic sources rich in Pi for subsequent use during germination and ensuing growth.

As for genes potentially involved in meiosis, the increased expression of dmc1, spo11, and some genes encoding MEI2-like proteins were found in dormant cysts of S. acuminata (Deng et al., 2017). At the same time, several mei2-like genes were downregulated. Nevertheless, the phase status — vegetative or sexual — has not been entirely clear for cysts in that case (Deng et al., 2017). Similar genes, namely spo11, dmc1, and also hop2, mnd1, msh4 and numerous mei2 homologs, are expressed at an elevated level during germination of S. acuminata dormant cysts formed in culture system, i.e., during the process when meiosis reliably occurs (Lin et al., 2022).

Simultaneous nitrogen and phosphorus deficiency can also trigger life cycle transitions, as expected. In some cases, combined nitrogen and phosphorus limitation produces the same effect as single-nutrient limitation. This has been described in Parvodinium cunningtonii, Peridinium cinctum, and Scrippsiella acuminata (Sako et al., 1984; Grigorszky et al., 2006; Yue et al., 2022) (Table S1). In other cases, combined deficiency can enhance the impact, for example, gamete fusion and planozygote encystment occurred in a larger proportion in -P-N medium than under N- or P-only deficient conditions in Gymnodinium catenatum (Figueroa et al., 2006b). The overview of nutrient deficiency-driven transitions between dinoflagellate life-cycle stages and associated changes in the expression of genes involved in meiotic process is presented in Figure 2.

Culture aging without direct altering in the medium composition also acts as conditions triggering life cycle switches. Although in this case it is more accurate to consider a combination of factors affecting the cell population, one of them will most likely be a deficiency of nutrients. Gamete pairs were most easily found in older cultures of Pfiesteria piscicida Steidinger & J.M.Burkholder (Parrow, Burkholder, 2004). Mating gametes occurred in the late exponential or stationary cultures of Prorocentrum cordatum, Prorocentrum donghaiense D.Lu and Karenia mikimotoii (Berdieva et al., 2020; Liu et al., 2020; Hu et al., 2022).

However, transition in the life cycle is a very complex process, which is regulated not only by simple nutrient deprivation but also by multiple physical and chemical factors, such as salinity, temperature and light intensity; and this effect was well demonstrated on Alexandrium minutum (Figueroa et al., 2011; Sixto et al., 2024). Decreased salinity and increased temperature together with phosphorus deprivation promoted planozygote formation and subsequent resting cysts production. In addition, specific irradiance level was shown to be an important factor promoting resting cysts production together with nutrient deficiency. For A. minutum, the optimal light intensity was 150 μE m⁻² s⁻¹. Higher irradiances reduced the total encystment yield (Sixto et al., 2024). The low nitrogen in combination with high irradiance had a significant impact on Peridinium cinctum (Chapman, Pfester, 1995). Specifically, in P. cinctum culture, only the highest temperature tested (22 °C) induced higher cyst yield under phosphorus deprivation compared with the control (full medium) (Grigorszky et al., 2006). Cell density is another important factor, which increases the probability of gamete mating. Moreover, presence of appropriate genetically diverse mating partners is a crucial factor for heterothallic species (Figueroa et al., 2011; Sixto et al., 2024).

7. Field Transcriptomic Evidence of Dinoflagellate Life Cycle Switching

Field transcriptomics is now intended to give direct, though still taxonomically specific, evidence that dinoflagellate life-cycle transitions leave a measurable gene-expression footprint in natural populations. In part, these footprints can be primarily used to detect and predict bloom development. Increased expression of genes associated with meiosis, such as dmc1, spo11, mnd1, msh6, rad21, rad51, exo1, mns1, smc3, smc2, and smc1, was demonstrated in metatranscriptome analysis of samples taken from natural populations of Prorocentrum shikokuense during the bloom (Lin et al., 2022). In Karenia mikimotoi bloom, the genes rad54, msh2, rad21, spo11, dmc1, mnd1, smc3 were upregulated. This observation became the basis of the “sex-for-proliferation” hypothesis, suggesting that sexual reproduction can facilitate population expansion or bloom maintenance without encystment (Lin et al., 2022). A recent study confirmed this hypothesis by showing the increase in gamete fusion and meiotic divisions (not encystment) during Dinophysis acuminata bloom peak (Sung-Clarke et al., 2026).

Yu et al. (2023) argued that bloom development and collapse of Karenia longicanalis Z.B.Yang, I.J.Hodgkiss & Gert Hansen were governed by multiple processes, especially energy and nutrient acquisition on the one hand and grazing, microbial attack, and community turnover – on the other. They have also identified 270 bloom-associated genes that included cell proliferation genes comprising those regulating sexual reproduction and cell division cycle (Yu et al., 2023). This shows the environmental context in which reproductive switching becomes visible: these are not isolated expression events in culture, but parts of a whole-community bloom trajectory. Three years of metatranscriptome analysis of Alexandrium minutum blooms showed that the species displayed a very specific expression pattern during natural blooms, with lower expression of photosynthesis and central-metabolism transcripts than the surrounding community (Metegnier et al., 2020). Moreover, despite the correlation of expression patterns with concentration of nutrients (phosphorus, nitrogen, silicate), genes coding for proteins involved in nutrient uptake and metabolism did not display extensive alterations (Metegnier et al., 2020).

Deng et al. (2025) used a dinoflagellate-specific DinoSL mRNA “hook” to analyze sediment-buried cyst assemblages and found that most major metabolic and regulatory pathways remained transcriptionally active, except photosynthesis; they also implicated autophagy and abscisic acid/gibberellin signaling in cyst dormancy and germination. That makes the sediment bank look less like a silent reservoir and more like a transcriptionally regulated dormant stage. Related sediment studies found Hsp90 and small heat-shock-protein transcripts in field-collected or buried cysts (Deng et al., 2021).

Culture-based studies make the field patterns easier to interpret because they identify the transcript modules that track encystment and dormancy. In Scrippsiella acuminata, encystment and cyst dormancy were associated with strong reprogramming of photosynthesis, active catabolism, meiotic recombination genes, MEI2-like genes, and abscisic-acid metabolism (Deng et al., 2017). In the later study, it has been demonstrated that energy storage dominated the immature-cyst stage, energy consumption dominated germination, and “genetic information processing” marked S. acuminata resting cyst (Wu et al., 2022). In Prorocentrum cordatum, cold-induced temporary cysts showed repression of calcium and protein kinase signaling, and some specific RNA-binding proteins, particularly proteins containing RNA-binding pentatricopeptide repeats and several MEI2-like proteins that have been determined as relevant to non-sexual cyst formation (Berdieva et al., 2025). These data still need to be summarized and processed to identify genes that could serve as molecular cues of processes occurring in natural populations. Such attempts have been repeatedly made for a long time. For example, Coyne and Cary (2005) showed that specific cyst transcripts could be detected both in laboratory cultures and directly in natural estuarine sediments by RT-PCR, supporting mRNA as a proxy for viable cysts.

8. Outlook: Molecular Biology Tools and Ecological Modeling Secure the Progress of Translational Aquatic Ecology

The expansion of HABs as a consequence of the enhanced inflow of anthropogenic nitrogen and phosphorus to aquatic ecosystems is one of the most crucial environmental problems, which causes detrimental economic consequences (Howarth, 2008; Gobler, 2020; Hallegraeff et al., 2021; Karlson et al., 2021; Orlova et al., 2022; Dai et al., 2023; Li et al., 2026; Telesh et al., 2026). Extensive studies of dinoflagellate blooms over the past decades have allowed identifying several important milestones on the way toward predicting red tides. These steps include conceptual and mathematical modeling based on the knowledge of molecular characteristics (e.g., regulation of cell cycle shifts and nutrients acquisition modes), individual cellular features (such as kleptoplasty efficiency, ability to form cysts and specific cysts dynamics), and major autecology characteristics (intra-population heterogeneity, auto-/heterotrophy balance, ecological niche sizes, etc.) of common HAB-forming dinoflagellate species (Figure 3).

Consideration of molecular mechanisms regulating life cycles, cysts production and excystment in these protists is essential for the development of basic ecological theories and practices. Among the latter, the theoretical basics of the classical intermediate disturbance hypothesis (Connell, 1978) and the resource competition theory (Tilman, 1977), as well as the newly suggested concepts like the protistan species maximum for the zone of critical salinity 5–8 (Telesh et al., 2011) and the mixoplankton concept (Flynn et al., 2019), are of particular importance for HABs modeling. The hypothesis describing the “sex-for-proliferation” scenario (as an alternative to “sex-for-encystment” concept) proposed by Lin with co-authors (2022) also expands the understanding of bloom development strategies. Genomics, transcriptomics, and molecular phylogeny data can substantially contribute to the progress of chaos theory (Benincà et al., 2008; Telesh et al., 2019; Cagle, Roelke, 2024) and secure the improvement of translational aquatic ecology in general (Skarlato et al., 2018). Coupling of the achievements of these theoretical and experimental studies with omics data and mathematical modeling is a basic prerequisite for the adequate comprehension and interpretation of the complex, nonlinear biotic interactions within aquatic communities and ecosystems.

Modeling studies currently span multiple levels of abstraction (Glibert et al., 2010; Horemans et al., 2023; Adamovich et al., 2025; Telesh et al., 2025b; Calbet, 2026). Trait-based models predict elevated trophic energy transfer efficiency, larger average organism size, and altered vertical carbon flux when mixotrophy is properly parameterized (Flynn, Mitra, 2009; Ward, Follows, 2016). Empirical habitat models can forecast bloom probability of dinoflagellates from certain physical and biogeochemical predictors (Horemans et al., 2023). The recent kleptoplasty model proposes the criterion for quantifying the contribution of plastids, retained by dinoflagellates during mixotrophy, to their photosynthesis, cells growth, and the autotrophy–heterotrophy balance (Telesh et al., 2025b).

Until recently, molecular studies of bloom-forming microorganisms and ecological research of the reasons and triggers, onset conditions, drivers of processing, causes of decay, and ecological consequences of algal blooms have been progressing largely in parallel. There has been little or no interference in their methodologies, although the studies aimed at resolving similar tasks like assessment of possible consequences of HABs. Now it has become possible to take an effort of merging the approaches of these disciplines within the framework of translational aquatic ecology, which implies close linkage of cellular biology, molecular ecology and omics with the classical ecological theories and practices (Skarlato et al., 2018). Application of the TAE strategy could help produce effective forecasts and thus come closer to combatting the devastating effects of HABs that enhance worldwide.

In this review, we attempted to unveil the linkages between the results of cellular and molecular biology studies of HAB-forming mixotrophic dinoflagellates with assessment of nutrient availability as a major modulator of their trophic strategies, diversity and life cycle regulation. In particular, it was shown that field transcriptomics likely provides direct evidence that dinoflagellate life-cycle transitions leave measurable gene-expression footprints in natural populations. These footprints, in turn, can be used in mathematical models to detect and predict the enhanced protist population growth and development leading to HABs. Likewise, a new ecological indicator, the kleptoplasty index, was developed because of conceptual and mathematical modeling, laboratory investigations of dinoflagellates physiology, and theoretical experiments. Coupling of these findings allowed assessing the quantitative contribution of kleptoplasty, an important intracellular process, to overall photosynthesis, cells proliferation, and finally – to population growth and the consequent HAB formation. The results highlighted in this review indicate that mathematical modeling can provide the primary opportunity to merge the newest outcomes of different disciplines – molecular biology and omics, protist cell morphology and physiology, field and laboratory experimental aquatic ecology, etc. – and thus secure a promising perspective for the mutually interlinked progress of these topical research directions.