Morphological variation and phylogeny of Karenia selliformis (Gymnodiniales, Dinophyceae) in an intensive cold-water algal bloom in eastern Hokkaido, Japan in September–November 2021

Harmful algal blooms responsible for mass mortalities of marine organisms have so far been rare in Hokkaido, northern Japan, although fish killing blooms have been frequently reported from western Japanese coasts. In September–November 2021, a huge and prolonged cold-water bloom occurred along the Pacific coast of eastern Hokkaido, Japan, and was associated with intensive mortalities of sea urchin, fish, octopus, shellfish, etc. In this study, morphology and phylogeny of the dominant and co-occurred unarmored dinoflagellates of the Kareniaceae in the bloom were examined by using light microscopy, scanning electron microscopy and molecular phylogeny inferred from ITS and LSU rDNA (D1–D3) sequences. Morphological observation and molecular phylogeny showed that the dominant species was Karenia selliformis, with co-occurrences of other kareniacean dinoflagellates, Kr. longicanalis, Kr. mikimotoi, Karlodinium sp., Takayama cf. acrotrocha, Takayama tuberculata and Takayama sp. The typical cell forms of K. selliformis in the bloom were discoid, dorsoventrally flattened, and larger than the cell sizes in previous reports, 35.3–43.6 (39.4±2.1) µm in length. Transparent cells of Kr. selliformis lacking or having several shrunken chloroplasts and oil droplets were also found. Cells of Kr. selliformis had morphological variation, but the species could be distinguished from other co-occurred Karenia species by its numerous (46–105) and small granular (2.9–4.6 µm in diameter) chloroplasts and the nucleus positioned in the hypocone. Cell density of Kr. selliformis exceeding 100 cells/mL was recorded in the range of temperature 9.8–17.6°C. The rDNA sequences determined from Kr. selliformis in the blooms of Hokkaido, Japan in 2021 were identical to those from another bloom in Kamchatka, Russia in 2020.

The Toxicity Testing of Cyanobacterial Toxins In Vivo and In Vitro by Mouse Bioassay: A Review

: Different biological methods based on bioactivity are available to detect cyanotoxins, including neurotoxicity, immunological interactions, hepatotoxicity, cytotoxicity, and enzymatic activity. The mouse bioassay is the first test employed in laboratory cultures, cell extracts, and water bloom materials to detect toxins. It is also used as a traditional method to estimate the LD50. Concerning the ease of access and low cost, it is the most common method for this purpose. In this method, a sample is injected intraperitoneally into adult mice, and accordingly, they are assayed and monitored for about 24 hours for toxic symptoms. The toxin can be detected using this method from minutes to a few hours; its type, e.g., hepatotoxin, neurotoxin, etc., can also be determined. However, this method is nonspecific, fails to detect low amounts, and cannot distinguish between homologues. Although the mouse bioassay is gradually replaced with new chemical and immunological methods, it is still the main technique to detect the bioactivity and efficacy of cyanotoxins using LD50 determined based on the survival time of animals exposed to the toxin. In addition, some countries oppose animal use in toxicity studies. However, high cost, ethical considerations, low-sensitivity, non-specificity, and prolonged processes persuade researchers to employ chemical and functional analysis techniques. The qualitative and quantitative analyses, as well as high specificity and sensitivity, are among the advantages of cytotoxicity tests to investigate cyanotoxins. The present study aimed at reviewing the results obtained from in-vitro and in-vivo investigations of the mouse bioassay to detect cyanotoxins, including microcystins, cylindrospermopsin, saxitoxins, etc.

PHYTOPLANKTON STATE AND CYANOTOXINS IN THE SVYATOZERO LAKE BLOOM SPOT (ONEGA LAKE BASIN, RUSSIA)

Introduction. This paper is the first to address the state of phytoplankton in eutrophic Svyatozero Lake (61о32´ N, 33о35´ E.), used for trout farming in the Republic of Karelia, in the North-Western Region of Russia, during the period of water bloom. For northern reservoirs, water bloom is not a typical phenomenon. However, due to climate warming and the large-scale development of trout breeding in the region, it becomes more urgent to study blooming reservoirs and related consequences for northern aquatic ecosystems and humans. Methods. We processed phytoplankton samples and measured photosynthesis using conventional methods. The concentration of chlorophyll a in water was determined spectrophotometrically, and the content of cyanotoxins was estimated by liquid chromatography-mass spectrometry. Results. In September 2019, we studied the structural, quantitative and functional characteristics of phytoplankton in the bloom spot. The phytoplankton abundance was 198.712 mln cells/l, the biomass was 14.945 mg/l, and the concentration of chlorophyll a reached 215.3 μg/l, which corresponded to the β-eutrophic state of the ecosystem in the study area. It was revealed that cyanobacteria corresponded to 99.8% of the biomass and 96.7% of the abundance. The species of the genus Microcystis were dominant (42%). They are well-known potential producers of cyanobacterial hepatotoxins, in particular, microcystins. We established the presence of microcystins. Their intracellular and extracellular concentrations were equal and in total amounted to 12.56 μg/l. We also identified eight structural variants of microcystins; among those, [D-Asp3] MC-RR accounted for up to 90% of the total content. The most toxic MC-LR variant was present in trace amounts only in biomass. According to the WHO standards, the content of microcystins and the number of cyanobacterial cells in the Svyatozero Lake bloom spot corresponds to average danger in the case of recreational use and can pose a threat to human health. Conclusion. It is necessary to monitor the phytoplankton composition, the presence of potentially toxic cyanobacterial species and cyanotoxins, as well as the state of trout farmed in the waters of Svyatozero Lake, since cyanotoxins can accumulate in fish tissues.

Vegetative cells may perform nitrogen fixation function under nitrogen deprivation in Anabaena sp. strain PCC 7120 based on genome-wide differential expression analysis

Nitrogen assimilation is strictly regulated in cyanobacteria. In an inorganic nitrogen-deficient environment, some vegetative cells of the cyanobacterium Anabaena differentiate into heterocysts. We assessed the photosynthesis and nitrogen-fixing capacities of heterocysts and vegetative cells, respectively, at the transcriptome level. RNA extracted from nitrogen-replete vegetative cells (NVs), nitrogen-deprived vegetative cells (NDVs), and nitrogen-deprived heterocysts (NDHs) in Anabaena sp. strain PCC 7120 was evaluated by transcriptome sequencing. Paired comparisons of NVs vs. NDHs, NVs vs. NDVs, and NDVs vs. NDHs revealed 2,044 differentially expressed genes (DEGs). Kyoto Encyclopedia of Genes and Genomes enrichment analysis of the DEGs showed that carbon fixation in photosynthetic organisms and several nitrogen metabolism-related pathways were significantly enriched. Synthesis of Gvp (Gas vesicle synthesis protein gene) in NVs was blocked by nitrogen deprivation, which may cause Anabaena cells to sink and promote nitrogen fixation under anaerobic conditions; in contrast, heterocysts may perform photosynthesis under nitrogen deprivation conditions, whereas the nitrogen fixation capability of vegetative cells was promoted by nitrogen deprivation. Immunofluorescence analysis of nitrogenase iron protein suggested that the nitrogen fixation capability of vegetative cells was promoted by nitrogen deprivation. Our findings provide insight into the molecular mechanisms underlying nitrogen fixation and photosynthesis in vegetative cells and heterocysts at the transcriptome level. This study provides a foundation for further functional verification of heterocyst growth, differentiation, and water bloom control.

FREE-LIVING CILIATES DURING THE PERIOD OF THE GREATEST CYANOBACTERIAL WATER BLOOM IN THE USINSKY BAY (KUIBYSHEV RESERVOIR)

The study of free-living ciliates in the Usinsky bay (the Kuibyshev reservoir) during the period of mass development of cyanobacteria are presented. The diversity of the ciliate species structure and spatial distribution were studied in summer. The abundance of ciliates varied from 124 to 1176 103ind./m3, biomass – from 6.9 mg/m3 to 104.9 mg/m3 during the study period. No significant differences in the communities of ciliates in different ecotopes of the bay (riverbed, open and overgrown by macrophytes coastal areas) were found. However, on average, in the riverbed part, all indicators of species richness and diversity, and quantitative developmentwere higher than in the littoral zone. The massive development of representatives of s/cl. Peritrichia (up to 51% of abundance and 66% of biomass): single cells species associated with planktonic algae (genera Vorticella, Vaginicola), and colonial free-floating Epistylisprocumbens Zacharias, 1897, was the distinctive feature of ciliate community in the UsinskyBay during the period of greatest cyanobacterial bloom. Water bloom significantly changed the trophic structure of the ciliate community: the role of algophages and nonselective omnivores, ordinary trophic groups of the Reservoir was significantly reduced, and bacterio-detritophages became absolutely dominated. It was shown that the dominant species of cyanobacteria had a significant effect on the ciliate community during the cyanobacterial water bloom.

Analysis and Diagnosis of Water Environment Pollution Sources of Tonggu Lake in Fuling District, Chongqing City

As an important urban lake in Fuling District, Chongqing City, there is water blooms in Tonggu Lake, which affect the ecological environment. This study analyzed and studied the external and internal status of water pollution through site survey, water quality testing and analysis, and laboratory data analysis, and diagnosed the water environment of Tonggu Lake, which is helpful for providing strong data support for the follow-up solution to the problem of water bloom in Tonggu Lake.

Monitoring of pollution factor of solidified body leachate in drilling well site and its influence on surface water environment

Aiming at the pollution of the leaching solution of the solidified body in the wellbore, a water eutrophication level evaluation algorithm based on the optimized analytic hierarchy process is proposed from the current situation of many factors affecting the eutrophication of the water body and the difficulty of evaluation. Based on this, the user-oriented PC software monitoring system was developed, which mainly includes three major modules: surface water environment monitoring, water eutrophication evaluation, water bloom prediction and warning, and system assistance. The surface water environment monitoring module can receive and display the surface water environment parameter information in real time, and has the functions of data comparison analysis and historical data search, and the eutrophication assessment and water bloom prediction and warning module can analyze the eutrophication status of water bodies in real time and provide short-term and medium-term warnings for water blooms. The research can promote the control of the leachate pollution of the wellbore in the wellbore and the optimization of the surface water environment.