Responses of Bloom-Forming Heterosigma akashiwo to Allelochemical Linoleic Acid: Growth Inhibition, Oxidative Stress and Apoptosis

Algal blooms have been occurring in many regions worldwide, and allelochemicals are important algaecides used to control harmful algal blooms (HABs). The allelopathic effects of linoleic acid (LA) on the harmful raphidophyte Heterosigma akashiwo were studied, and the possible mechanisms were investigated through analyses of population growth dynamics, cellular ultrastructure and the physiological levels of H. akashiwo. The results showed that the inhibitory effect of LA on H. akashiwo cells increased with an increasing LA concentration. The levels of ROS and MDA were significantly elevated, indicating oxidative stress and lipid peroxidation due to LA exposure. At the same time, LA also activated the antioxidant system, including superoxide dismutase (SOD), catalase (CAT), and POD, and non-enzymatic antioxidants such as reduced AsA and glutathione (GSH). Transmission electron microscopy (TEM) revealed that the morphology of the algal cells was impaired in an LA-dependent manner. Annexin V-FITC/PI double staining and flow cytometric analysis revealed that LA exposure decreased the cellular mitochondrial membrane potential (MMP), increased the rate of apoptosis. LA modulated bcl-2/bax homeostasis and increased the expressions of cytochrome c and caspases-3 and -9, proving that LA induced cell death via the mitochondria-mediated apoptotic pathway. It was suggested that LA had allelopathic effects on H. akashiwo, inducing physiological and morphological changes and finally triggering the apoptosis of H. akashiwo. All of these results showed that LA might have the potential as an algaecide to control harmful algae.

De novo assembly and inferred functional annotation of the transcriptome of Heterosigma akashiwo

Heterosigma akashiwo is a eukaryotic, cosmopolitan, and unicellular alga (class: Raphidophyceae), and produces fish-killing blooms. There is a substantial scientific and practical interest in its ecophysiological characteristics that determine bloom dynamics and its adaptation to broad climate zones. A well-annotated genomic/genetic sequence information enables researchers to characterize organisms using modern molecular technology. The Chloroplast and the mitochondrial genome sequences and transcriptome sequence assembly (TSA) datasets with limited sizes for H. akashiwo are available in NCBI nucleotide database on December 2021: there is no doubt that more genetic information of the species will greatly enhance the progress of biological characterization of the species. Here, we conducted H. akashiwo RNA sequencing, a de novo transcriptome assembly (NCBI TSA ICRV01) of a large number of high-quality short-read sequences, and the functional annotation of predicted genes. Based on our transcriptome, we confirmed that the organism possesses genes that were predicted to function in phagocytosis, supporting the earlier observations of H. akashiwo bacterivory. Along with its capability for photosynthesis, the mixotrophy of H. akashiwo may partially explain its high adaptability to various environmental conditions. Our study here will provide an important toehold to decipher H. akashiwo ecophysiology at a molecular level.

De novo assembly and inferred functional annotation of the transcriptome of Heterosigma akashiwo

Heterosigma akashiwo is a eukaryotic, cosmopolitan, and unicellular alga (class: Raphidophyceae), and produces fish-killing blooms. There is a substantial scientific and practical interest in its ecophysiological characteristics that determine bloom dynamics and its adaptation to broad climate zones. A well-annotated genomic/genetic sequence information enables researchers to characterize organisms using modern molecular technology. The Chloroplast and the mitochondrial genome sequences and transcriptome sequence assembly (TSA) datasets with limited sizes for H. akashiwo are available in NCBI nucleotide database on December 2021: there is no doubt that more genetic information of the species will greatly enhance the progress of biological characterization of the species. Here, we conducted H. akashiwo RNA sequencing, a de novo transcriptome assembly (NCBI TSA ICRV01) of a large number of high-quality short-read sequences, and the functional annotation of predicted genes. Based on our transcriptome, we confirmed that the organism possesses genes that were predicted to function in phagocytosis, supporting the earlier observations of H. akashiwo bacterivory. Along with its capability for photosynthesis, the mixotrophy of H. akashiwo may partially explain its high adaptability to various environmental conditions. Our study here will provide an important toehold to decipher H. akashiwo ecophysiology at a molecular level.

Growth and Nutrient Uptake Characteristics of Heterosigma akashiwo (Raphidophyceae) under Nitrogen and Phosphorus Concentrations in the East China Sea

Heterosigma akashiwo is classified as a harmful algal bloom (HAB) species that frequently occurs in eutrophic coastal waters and results in the contamination and mortality of fish and shellfish. The growth of H. akashiwo in four phosphate and nitrate concentration scenarios, representing the observed nutrient concentration ranges in the East China Sea (ECS), was evaluated to further understand the effect of nutrient concentrations on H. akashiwo blooms. The specific growth rate in the exponential growth phase (µ′) and the maximum cell density were lower (17–21% and 41%, respectively) under low phosphorus concentration scenarios, compared to the rates observed under high phosphorus concentration scenarios. The cellular nitrogen-to-phosphorus ratios of H. akashiwo were influenced by the initially supplied N:P ratio and the allocation strategy employed. Phosphorus concentration had a greater influence on the total growth of H. akashiwo than nitrate did, within the natural nutrient conditions of the ECS. These results could serve as a reference for coastal water management and marine ecological management and may be useful for further studies on the simulation and prediction of H. akashiwo blooms, particularly in the ECS.

Toxicity of Naphthenic Acids on the Chlorophyll Fluorescence Parameters and Antioxidant Enzyme Activity of Heterosigma akashiwo

Petroleum hydrocarbons can serve as a carbon source for marine phytoplankton; so, marine high-acid crude oil pollution events are likely to result in algal outbreaks or harmful algal blooms (HABs) in surface waters. Naphthenic acids (NAs) are the primary acidic component of crude oil, and red tide is of great concern due to its high diffusivity and strong destructive properties. It is important to study the mechanism of the toxic effect of NAs on the typical red tide algae, Heterosigma akashiwo, for the balance and stability of marine algae. The mechanism of NAs’ damage effect was investigated in terms of the antioxidant enzyme activity, cell number, the chlorophyll positive fluorescence parameters, and the cell morphology of microalgae. Experiments confirmed the hormesis of low-concentration (0.5, 2, and 4 mg/L) NAs on Heterosigma akashiwo, and the indicators of high-concentration (8 and 16 mg/L) NA exposures showed inhibition. In this study, the toxic effect of NAs on the target organism showed a clear concentration–dose relationship. The 16 mg/L NAs stress caused severe damage to the morphology and structure of the target biological cells in a short time (96 h), and the population growth decreased. The target organisms showed a staged oxidative stress response to NAs. The behavior in the low-concentration treatment groups showed toxicant excitatory effects on the photosynthetic efficiency and antioxidant enzyme activity of the target organisms. This study provides theoretical and practical data for the development of an important toxicological model of the toxicant’s excitement effects and antioxidant defense mechanisms. In addition, it provides prospective research data for the prediction and avoidance of ecological risk from NA pollution in marine environments.