Freshwater Strains Research Articles

Proteins Associated with Salinity Adaptation of the Dinoflagellates: Diversity and Potential Involvement in Species Evolution

Protists inhabit marine, brackish and fresh waters. The salt barrier plays an important role in the origin of their diversity. Salinity tolerance differs among species and sometimes even among different strains of the same species, indicating local adaptation. Dinoflagellates from the Apocalathium genus are represented by at least four species, which originated via rapid and recent radiation. Water salinity was suggested as one of the key factors for this radiation. A previous study found RNA transcripts, which belong exclusively to saline strains of Apocalathium, and were absent in its freshwater strains. In the present paper, the diversity of these transcripts and their orthologs from marine and freshwater protists were analysed using bioinformatic approaches. First, it was found that these specific transcripts translated to the proteins, which are important for osmoregulation (e.g., transport of various compounds including glycine betaine, regulation of microtubule organisation, post transcriptional modifications). This supports the idea that speciation within Apocalathium resulted in the loss of osmoregulatory genes by freshwater species. Second, protein distribution was not highly species specific, because their orthologs were found in different dinoflagellates and were relatively common in other phototrophic protists, though the sequences were highly variable. Proteins from 13 orthogroups were absent or very rare in studied freshwater genomes and transcriptomes. They could play a specific role in protists salinity tolerance. Third, detailed phylogenetic analyses of betaine-like transporter and chloride transmembrane transporters, which probably are one of the key proteins associated with salinity tolerance, revealed high levels of multiple and variable copies that were not eliminated from the genome during the evolution. The expression of their genes could be important in the adaptation of dinoflagellates to salinity changes, as it was already shown for some other protists.

Ploidy levels in diverse picocyanobacteria from the Baltic Sea.

In nature, the number of genome or chromosome copies within cells (ploidy) can vary between species and environmental conditions, potentially influencing how organisms adapt to changing environments. Although ploidy levels cannot be easily determined by standard genome sequencing, understanding ploidy is crucial for the quantitative interpretation of molecular data. Cyanobacteria are known to contain haploid, oligoploid, and polyploid species. The smallest cyanobacteria, picocyanobacteria (less than 2 μm in diameter), have a widespread distribution ranging from marine to freshwater environments, contributing significantly to global primary production. In this study, we determined the ploidy level of genetically and physiologically diverse brackish picocyanobacteria isolated from the Baltic Sea using a qPCR assay targeting the rbcL gene. The strains contained one to four genome copies per cell. The ploidy level was not linked with phylogeny based on the identity of the 16S rRNA gene. The variation of ploidy among the brackish strains was lower compared to what has been reported for freshwater strains and was more similar to what has been reported for marine strains. The potential ecological advantage of polyploidy among picocyanobacteria has yet to be described. Our study highlights the importance of considering ploidy to interpret the abundance and adaptation of brackish picocyanobacteria.

Cyanoleptonema yuqiaoensis gen. & sp. nov. (Oscillatoriaceae, Oscillatoriales), a novel filamentous cyanobacterium isolated from a eutrophic reservoir in China

ABSTRACT An unknown filamentous cyanobacterium was isolated from the Yuqiao Reservoir, a eutrophic drinking water reservoir in Tianjin city, northern China. The cyanobacterium shares some morphological similarities with most Leptolyngbya-like species, having thin trichomes that can facultatively produce sheaths, with one trichome per sheath, and contain peripheral thylakoids. Based on a combination of morphological, molecular, ultrastructural and ecological evidence, we propose the description of a new cyanobacterial taxon, Cyanoleptonema yuqiaoensis gen. & sp. nov. which expands our understanding of cyanobacterial diversity. Phylogenetic analyses based on 16S rRNA gene sequences placed this freshwater strain into a well-supported clade closely related to the marine or saline genera Sodalinema and Baaleninema. Genetic identity between Cyanoleptonema and other cyanobacteria, including Sodalinema and Baaleninema, was found to be less than 95% in 16S rRNA. The genus was further distinguished by morphological and ecological characteristics, including cell size and the ratio between cell length and width.

Optimizing biofuel production from algae using four-element framework: Insights for maximum economic returns

This paper introduces a Four Element Strategy (FES) focused on concentration of CO2 gas (Cc,g), energy input to the process, concentration of nutrients, and the type of algal strain to maximize biofuel production (BYP) from algae strains and evaluate associated economic benefits. Two algal strains encompassing slain algae Nannochloropsis sp. (Ns), and freshwater strain Chlorella vulgaris (Cv) were used in the current study. Response Surface Methodology (RSM) and Box Behnkin Design (BBD) are integrated with a biokinetic model to explore the relationships among growth parameters in the ranges; Cc,g ∼ 0.03–20 %), Nitrogen (N) to phosphorus (P) ratio (RN/P)∼1–11, and light intensity (LI)∼ 100–400 µE m⁻² s⁻¹ with corresponding CO2 bio-fixation rate (RCO2, g L⁻¹ d⁻¹), biomass accumulation (X, g L⁻¹), and algal growth rate (µ, d⁻¹). Optimization results indicate that the most productive conditions involve LI in the range of 100–270 µE m⁻² s⁻¹, Cc,g between 0.03 % and 15 %, and RN/P of 3.5–11, achieving maximum RCO2, X, and µ at 1 g L⁻¹ d⁻¹, 6 g L⁻¹, and 1.6 d⁻¹, respectively. Further optimization using the desirability function supports achieving RCO2 of 1.2 g L⁻¹ d⁻¹, X of 1.8 g L⁻¹, and µ of 1.7 d⁻¹, with RN/P, Cc,g, and LI at 11, 4.5 %, and 200 µE m⁻² s⁻¹, respectively. Notably, Ns demonstrated higher nutrient uptake rates and BYP compared to Cv, with total production costs of $0.629/L and respective required cost coverages of $0.138 and $0.375 for Ns and Cv. This study offers a comprehensive guideline for large-scale microalgae cultivation technology (MCT) processes, providing insights into optimizing conditions for biofuel production while considering economic viability.

Towards bioprocess engineering of cable bacteria: Establishment of a synthetic sediment.

Cable bacteria, characterized by their multicellular filamentous growth, are prevalent in both freshwater and marine sediments. They possess the unique ability to transport electrons over distances of centimeters. Coupled with their capacity to fix CO2 and their record-breaking conductivity for biological materials, these bacteria present promising prospects for bioprocess engineering, including potential electrochemical applications. However, the cultivation of cable bacteria has been limited to their natural sediment, constraining their utility in production processes. To address this, our study designs synthetic sediment, drawing on ion exchange chromatography data from natural sediments and existing literature on the requirements of cable bacteria. We examined the effects of varying bentonite concentrations on water retention and the impacts of different sands. For the first time, we cultivated cable bacteria on synthetic sediment, specifically the freshwater strain Electronema aureum GS. This cultivation was conducted over 10 weeks in a specially developed sediment bioreactor, resulting in an increased density of cable bacteria in the sediment and growth up to a depth of 5 cm. The creation of this synthetic sediment paves the way for the reproducible cultivation of cable bacteria. It also opens up possibilities for future process scale-up using readily available components. This advancement holds significant implications for the broader field of bioprocess engineering.

Nitrite Degradation by a Novel Marine Bacterial Strain Pseudomonas aeruginosa DM6: Characterization and Metabolic Pathway Analysis

High concentrations of nitrite in marine aquaculture wastewater not only pose a threat to the survival and immune systems of aquatic organisms but also contribute to eutrophication, thereby impacting the balance of coastal ecosystems. Compared to traditional physical and chemical methods, utilizing microorganism-mediated biological denitrification is a cost-effective and efficient solution. However, the osmotic pressure changes and salt-induced enzyme precipitation in high-salinity seawater aquaculture environments may inhibit the growth and metabolism of freshwater bacterial strains, making it more suitable to select salt-tolerant marine microorganisms for treating nitrite in marine aquaculture wastewater. In this study, a salt-tolerant nitrite-degrading bacterium, designated as DM6, was isolated from the seawater (salinity of 25–30‰) of Portunus trituberculatus cultivation. The molecular identification of strain DM6 was conducted using 16S rRNA gene sequencing technology. The impacts of various environmental factors on the nitrite degradation performance of strain DM6 were investigated through single-factor and orthogonal experiments, with the selected conditions considered to be the key factors affecting the denitrification efficiency of microorganisms in actual wastewater treatment. PCR amplification of key genes involved in the nitrite metabolism pathway of strain DM6 was conducted, including denitrification pathway-related genes narG, narH, narI, nirS, and norB, as well as assimilation pathway-related genes nasC, nasD, nasE, glnA, gltB, gltD, gdhB, and gdhA. The findings indicated that strain DM6 is classified as Pseudomonas aeruginosa and exhibits efficient nitrite degradation even under a salinity of 35‰. The optimal nitrite degradation efficiency of DM6 was observed when using sodium citrate as the carbon source, a C/N ratio of 20, a salinity of 13‰, pH 8.0, and a temperature of 35 °C. Under these conditions, DM6 could completely degrade an initial nitrite concentration of 156.33 ± 1.17 mg/L within 36 h. Additionally, the successful amplification of key genes involved in the nitrite denitrification and assimilation pathways suggests that strain DM6 may possess both denitrification and assimilation pathways for nitrite degradation simultaneously. Compared to freshwater strains, strain DM6 demonstrates higher salt tolerance and exhibits strong nitrite degradation capability even at high concentrations. However, it may be more suitable for application in the treatment of wastewater from marine aquaculture systems during summer, high-temperature, or moderately alkaline conditions.

Chlorine Photolysis: A Step Forward in Inactivating Acanthamoeba and Their Endosymbiont Bacteria

Chlorine and solar disinfection are widely used disinfectants in water treatment. However, certain potential pathogens can resist these methods, posing a public health risk. One such case is Acanthamoeba, a resistant free-living amoeba that protects pathogens inside from disinfection, thus endangering the health of water users. This work is the first evaluation of the inactivation efficiency achieved by combining NaClO (Cl2) and solar radiation (SR) against two Acanthamoeba strains from different sources (freshwater and pool water) and their endosymbiont bacteria (EB). Amoebae were exposed to different Cl2 doses (0–500 mg/L), SR wavelength ranges (280–800 nm and 320–800 nm), used as gold standards, and their combinations. The EB exhibited resistance to conventional Cl2 and SR treatments, requiring up to 20 times higher disinfectant doses than those needed to inactivate their protective Acanthamoeba. The pool strain and its EB demonstrated greater resistance to all treatments compared to the freshwater strain. Treatments with Cl2 (5 mg/L)/SR280–800nm completely inactivated both Acanthamoeba and EB of the freshwater strain, reducing up to 100 times the necessary Cl2 doses, suggesting that chlorine photolysis is an attractive treatment for disinfecting freshwater and preventing waterborne diseases associated with Acanthamoebae and its EB.

Fatty acids composition of diatoms from the genus Mayamaeya

Diatoms are the one of the most widespread and diverse groups of algae. However, even now the numbers of the works devoted to the study of their fatty acid composition isn’t enough. In this paper, the fatty acid profiles of soil and freshwater strains of the genus Mayamaea were described. Despite the fact that the strains were isolated from different ecosystems, minor differences in the compiled fatty acid profiles were noted. In the course of the work, it was revealed that saturated palmitic and stearic acids and monounsaturated palmitoleic acid are dominant for the studied objects.

Elucidating the picocyanobacteria salinity divide through ecogenomics of new freshwater isolates

BackgroundCyanobacteria are the major prokaryotic primary producers occupying a range of aquatic habitats worldwide that differ in levels of salinity, making them a group of interest to study one of the major unresolved conundrums in aquatic microbiology which is what distinguishes a marine microbe from a freshwater one? We address this question using ecogenomics of a group of picocyanobacteria (cluster 5) that have recently evolved to inhabit geographically disparate salinity niches. Our analysis is made possible by the sequencing of 58 new genomes from freshwater representatives of this group that are presented here, representing a 6-fold increase in the available genomic data.ResultsOverall, freshwater strains had larger genomes (≈2.9 Mb) and %GC content (≈64%) compared to brackish (2.69 Mb and 64%) and marine (2.5 Mb and 58.5%) isolates. Genomic novelties/differences across the salinity divide highlighted acidic proteomes and specific salt adaptation pathways in marine isolates (e.g., osmolytes/compatible solutes - glycine betaine/ggp/gpg/gmg clusters and glycerolipids glpK/glpA), while freshwater strains possessed distinct ion/potassium channels, permeases (aquaporin Z), fatty acid desaturases, and more neutral/basic proteomes. Sulfur, nitrogen, phosphorus, carbon (photosynthesis), or stress tolerance metabolism while showing distinct genomic footprints between habitats, e.g., different types of transporters, did not obviously translate into major functionality differences between environments. Brackish microbes show a mixture of marine (salt adaptation pathways) and freshwater features, highlighting their transitional nature.ConclusionsThe plethora of freshwater isolates provided here, in terms of trophic status preference and genetic diversity, exemplifies their ability to colonize ecologically diverse waters across the globe. Moreover, a trend towards larger and more flexible/adaptive genomes in freshwater picocyanobacteria may hint at a wider number of ecological niches in this environment compared to the relatively homogeneous marine system.

Insights from cyanobacterial genomic and transcriptomic analyses into adaptation strategies in terrestrial environments

Phylogenomic analysis of Nostocsp. MG11, a terrestrial cyanobacterium, and some terrestrial and freshwater Nostoc strains showed that the terrestrial strains grouped together in a distinctive clade, which reveals the effect of habitat on shaping Nostoc genomes. Terrestrial strains showed larger genomes and had higher predicted CDS contents than freshwater strains. Comparative genomic analysis demonstrated that genome expansion in the terrestrial Nostoc is supported by an increase in copy number of the core genes and acquisition of shared genes. Transcriptomic profiling analysis under desiccation stress revealed that Nostoc sp. MG11 protected its cell by induction of catalase, proteases, sucrose synthase, trehalose biosynthesis and maltodextrin utilization genes and maintained its normal metabolism during this condition by up-regulation of genes related to phycobilisomes and light reactions of photosynthesis, CO2 fixation and protein metabolism. These results provide insights into the strategies related to survival and adaptation of Nostoc strains to terrestrial environments.

Resistance of DISCOVR algae strains to deleterious species

The goal of the DISCOVR consortium is to improve the annualized areal production of open algal production systems through a process of algal strain evaluation and selection combined with testing in outdoor pilot scale production systems. The role of Sandia National Laboratories, in this evaluation process, has been to screen the selected subset of algal strains for resistance to grazing. Sandia tested 9 marine and 8 freshwater strains of algae against a panel of 14 grazer species, (5 marine, 5 freshwater and 4 euryhaline). A total of 153 algal/grazer pairs at two grazer concentrations for a total of 306 assays in triplicate. The relative resistance of each algal strain to each grazer strain was expressed as the decimal fraction of its specific growth rate 29in the presence of the grazer divided by the growth rate of the algae alone. Of the strains tested, those with the greatest resistance to the grazer panel were the freshwater strains Acutodesmus obliquus UTEX 393 and Scenedesmus obliquus. DOE 152z, and the marine strains Micractinium sp 14-F2 and Scenedesmus sp. 46B-D3. Additionally, these assays revealed the unexpected levels of variation in grazer resistance between taxonomically related organisms.

Biotransformation of phthalate plasticisers and bisphenol A by marine and freshwater fungi

Phthalate esters (PEs) are environmentally ubiquitous micropollutants that are used as plasticizers and additives in diverse consumer products. Considerable concern relates to their reported xenoestrogenicity and the microbial-based attenuation of environmental PE concentrations is of interest to combat harmful downstream effects. Fungal PE catabolism has received less attention than that by bacteria, and particularly marine fungal species remain largely overlooked in this respect. We have compared the biocatalytic and biosorptive removal rates of di-n-butyl phthalate (DBP) and diethyl phthalate (DEP), chosen as two environmentally prominent PE representatives (exhibiting differing structures and hydrophobicities), by marine- and freshwater fungal strains. Bisphenol A, both an extensively used plastic additive and prominent environmental xenoestrogen, was included as a reference compound due to its previously well-documented fungal degradation. Partial pathways for DBP metabolization by these ecophysiologically diverse ascomycetes were proposed with the help of UPLC-QTOF-MS analysis. Species-specific biochemical reaction steps contributing to DBP metabolism were also observed. The involved reactions include initial cytochrome P450-dependent monohydroxylations of DBP with subsequent further oxidation of related metabolites, de-esterification via either hydrolytic cleavage or cytochrome P450-dependent oxidative O-dealkylation, transesterification, and demethylation steps - finally yielding phthalic acid as a central intermediate in all pathways. Beyond previous research into fungal PE metabolism which emphasises hydrolytic de-esterification as the primary catabolic step, a prominent role of cytochrome P450 monooxygenase-catalysed reactions is established.

Characterisation of the volatile profile of microalgae and cyanobacteria using solid-phase microextraction followed by gas chromatography coupled to mass spectrometry

Microalgae and microalgae-derived ingredients are one of the top trends in the food industry. However, consumers’ acceptance and purchase intention of a product will be largely affected by odour and flavour. Surprisingly, the scientific literature present a very limited number of studies on the volatile composition of microalgae and cyanobacteria. In order to fill the gap, the main objective of the present study was to elucidate the volatile composition of seven microalgal and cyanobacterial strains from marine and freshwaters, with interest for the food industry while establishing its potential impact in odour. Among the seven selected strains, Arthrospira platensis showed the highest abundance and chemical diversity of volatile organic compounds (VOCs). Aldehydes, ketones, and alcohols were the families with the highest diversity of individual compounds, except in Arthrospira platensis and Scenedesmus almeriensis that showed a profile dominated by branched hydrocarbons. Marine strains presented a higher abundance of sulfur compounds than freshwater strains, while the ketones individual profile seemed to be more related to the taxonomical domain. The results of this study indicate that the VOCs composition is mainly driven by the individual strain although some volatile profile characteristics could be influenced by both environmental and taxonomical factors.

A Fast-Growing Oleaginous Strain of Coelastrella Capable of Astaxanthin and Canthaxanthin Accumulation in Phototrophy and Heterotrophy.

Considering the importance of microalgae as a promising feedstock for the production of both low- and high-value products, such as lipids and pigments, it is desirable to isolate strains which simultaneously accumulate these two types of products and grow in various conditions in order to widen their biotechnological applicability. A novel freshwater strain from the genus Coelastrella was isolated in Belgium. Compared to other Coelastrella species, the isolate presented rapid growth in phototrophy, dividing 3.5 times per day at a light intensity of 400 µmol·m−2·s−1 and 5% CO2. In addition, nitrogen depletion was associated with the accumulation of astaxanthin, canthaxanthin, and fatty acids, which reached ~30% of dry weight, and a majority of SFAs and MUFAs, which are good precursors for biodiesel. This strain also accumulated astaxanthin and canthaxanthin in heterotrophy. Although the content was very low in this latter condition, it is an interesting feature considering the biotechnological potential of the microalgal heterotrophic growth. Thus, due to its rapid growth in the light, its carotenogenesis, and its fatty acids characteristics, the newly identified Coelastrella strain could be considered as a potential candidate for biorefinery purposes of both low- and high-values products.

Comparative analysis of phycoerythrin production in cryptophytes

Phycobiliproteins are pigments with uses in pharmacology, cosmetics, foods, and as fluorescent probes in biochemistry. Cryptophyte microalgae are one possible source of phycobiliproteins as well as other molecules such as omega-3 fatty acids. The use of cryptophytes in biotechnology is currently very limited and especially the potential of freshwater species is poorly documented. For commercial microalgae production, it is important to find the best performing strains in terms of growth and yields of the products of interest. Phycoerythrin is a phycobiliprotein with red colour and strong yellow fluorescence. In this study, we evaluate the growth and phycoerythrin production of eight strains of freshwater cryptophytes belonging to the genus Cryptomonas, comparing them to two marine strains. The strains are grown in batch cultures under standardised conditions. Most of the studied freshwater strains have lower growth rates and all of them have lower biomass yields than the marine strains. However, most of them have much higher cellular phycoerythrin concentrations, which in the case of two strains leads them to a significantly higher overall phycoerythrin yield. There is large variation among cryptophytes in growth rates and phycoerythrin content. Our results suggest that freshwater cryptophytes of the genus Cryptomonas may be better sources of phycoerythrin than the more extensively studied marine strains.

Metabolic stability of freshwater Nitzschia palea strains under silicon stress associated with triacylglycerol accumulation

Si is an essential nutrient for diatom growth. It is involved in frustule formation and is directly linked to the cell cycle. Despite numerous studies analyzing Si starvation in diatoms, the interaction between Si and central metabolism remains poorly understood. Accordingly, we analyzed the metabolite profiles of two freshwater strains of Nitzschia palea, BR006 and BR022 (a fast and a slow-growing strain, respectively), under different Si concentrations (0, 0.25, 1.25, and 5 mM) in different harvesting points. The strains exhibited similar metabolic responses under Si starvation, such as high pigment and total lipid, as well as high primary metabolite accumulation; only ornithine level was low in both strains in the Si-starved condition. However, the response of each strain differed under different Si concentrations. Triacylglycerol (TAG) accumulation in cells was inversely proportional to growth rate. Unlike BR006, slow-growing BR022 cells exhibited high TAG production even under high Si conditions. These results suggest that under optimal Si conditions, in the fast-growing BR006 strain, carbon from photosynthesis is directed toward new cell formation, while in the slow-growing BR022 cells, carbon is stored as TAG as a priority.

Lipid accumulation by Coelastrella multistriata (Scenedesmaceae, Sphaeropleales) during nitrogen and phosphorus starvation

A novel freshwater strain of Coelastrella multistriata MZ–Ch23 was discovered in Tula region, Russia. The identification is based on morphological features, phylogenetic analysis of SSU rDNA gene and ITS1–5.8S rDNA–ITS2 region and predicted secondary structure of the ITS2. Phylogenetic analysis places the novel strain in the “core” Coelastrella clade within the Chlorophyceae. This is the first record of Coelastrella multistriata in the algal flora of Russia. Cultivation experiments were carried out to evaluate growth dynamics of the newly identified strain and the impact of nitrogen and/or phosphorus depletion on the fatty acid profiles and lipid productivity. On the fully supplemented Bold’s basal medium and under phosphorus-depleted conditions as well, the fatty acid profiles were dominated by α-linolenic acid (29.4–38.1% of total fatty acids). Depletion of either nitrogen or both nitrogen and phosphorus was associated with increased content of oleic acid (32.9–33.7%) and linoleic acid (11.9%). Prolongation of the growth to two months (instead of 25 days) resulted in increased content and diversity of very long-chain fatty acids including saturated species. The total very long-chain fatty acid content of 9.99% achieved in these experiments was 1.9–12.3-fold higher than in stress experiments. The highest variation was observed for oleic acid (3.4–33.7%). The novel strain showed the ability to accumulate lipids in amounts up to 639.8 mg L−1 under nitrogen and phosphorus starvation, which exceeds the previously obtained values for most Coelastrella strains. Thus, the newly identified MZ–Ch23 strain can be considered as a potential producer of omega-3 fatty acids on fully supplemented Bold’s basal medium or as a source of biomass with high content of saturated and monounsaturated fatty acids after nitrogen and phosphorus starvation.

Assessing the salt tolerance of Spirulina platensis freshwater strains and examining cheap culture media for cultivation of the potential strain

Spirulina cyanobacteria have been widely cultivated to exploit products such as crude protein, vitamins, phycocyanin pigment... with high nutritional and pharmacological values. However, the commercialization of these products is still a challenging issue due to high biomass cost, which is mainly caused by expensive nutrients in the culture medium. In this study, from 11 freshwater S. platensis strains, by culture screening, we found 7 strains being capable of profitable growth on inexpensive seawater with salinity ranging from 5 - 30‰, and selected ST strain as the potential strain for further study. Natural seawater must be pretreated to remove ions that easily cause precipitation of nutrients in the culture medium such as Mg2+, Ca2+, SO42-… before using. The ST strain showed the best growth in the natural seawater medium with 30‰ salinity containing 3 g/L NaNO3, 0.5 g/L K2HPO4, 0.05 g/L FeSO4. This strain reached the highest biomass yield at 0.487 g/L and the specific growth rate (µ) of 0.12 x day-1; protein and phycocyanin contents reached 48.6% and 127 mg/g of dry biomass, respectively. There was no difference in the mentioned above values with biological statistical significance between this medium and SOT medium in distilled water. The ST strain biomass was qualified to be used for the production of functional foods. Results of this study provided scientific basis for the use of marine and brackish waters to produce biomass of this highly economic cyanobacterium.

Experimental freezing of freshwater pennate diatoms from polar habitats.

Diatoms are microalgae that thrive in a range of habitats worldwide including polar areas. Remarkably, non-marine pennate diatoms do not create any morphologically distinct dormant stages that could help them to successfully face unfavourable conditions. Their survival is probably connected with the adaptation of vegetative cells to freezing and desiccation. Here we assessed the freezing tolerance of vegetative cells and vegetative-looking resting cells of 12 freshwater strains of benthic pennate diatoms isolated from polar habitats. To test the effect of various environmental factors, the strains were exposed to -20 °C freezing in four differently treated cultures: (1) vegetative cells growing in standard conditions in standard WC medium and (2) resting cells induced by cold and dark acclimation and resting cells, where (3) phosphorus or (4) nitrogen deficiency were used in addition to cold and dark acclimation. Tolerance was evaluated by measurement of basal cell fluorescence of chlorophyll and determination of physiological cell status using a multiparameter fluorescent staining. Four strains out of 12 were able to tolerate freezing in at least some of the treatments. The minority of cells appeared to be active immediately after thawing process, while most cells were inactive, injured or dead. Overall, the results showed a high sensitivity of vegetative and resting cells to freezing stress among strains originating from polar areas. However, the importance of resting cells for survival was emphasized by a slight but statistically significant increase of freezing tolerance of nutrient-depleted cells. Low numbers of surviving cells in our experimental setup could indicate their importance for the overwintering of diatom populations in harsh polar conditions.

The Complete Genome of Nocardia seriolae MH196537 and Intra-Species Level as Analyzed by Comparative Genomics Based on Random Forest Algorithm

Nocardiosis is a major problem affecting fish that are farmed in seacages as well as freshwater fish; therefore, deciphering the bacteriological features of Nocardia seriolae is crucial. In particular, a number of studies over the past two years have reported the genome sequence of N. seriolae, and a comparative genomics approach is expected to yield valuable information on its epidemiological characteristics. The purpose of this study was to perform whole-genome sequence analysis of N. seriolae MH196537 from the Japanese eel and to investigate the significant differences noted between strains isolated from freshwater fish and marine fish by using Random Forest, a reliable machine learning algorithm. The Pacbio platform was employed to sequence the MH196537 strain, and genomic information from the other 16 strains was used for comparative analyses. All coding sequences of the 17 strains were categorized in RASTtk Sub-systems. The MH196537 strain had one contig, and it shared a high average nucleotide identity (ANI) with the freshwater strains (0.9994 - 0.9999) rather than the seawater strains (0.9985 - 0.9994). Moreover, 22 RASTtk subsystems carried a different number of genes from each N. seriolae. The fatty acids, lipids, and isoprenoids subsystem showed the highest mean decrease in the Gini index of over 1.5. Interestingly, freshwater strains were found to harbor all of the genes for both the mevalonate (MVA) and non-mevalonate pathways (MEP), whereas only the MEP existed in strains from diseased marine fish. Considering the differences in the byproducts of isoprenoids from the different pathways, it is likely that this will affect host-pathogen interactions; therefore, harboring the different pathways for the synthesis of isoprenoids could be an important pathogenic factor of N. seriolae.