The generalized response of the cyanobacterium Nostoc sp. PCC 7120 to and its recovery from phosphorus (P) starvation stress differ drastically under diazotrophic and non-diazotrophic growth modes. In nitrogen (N) -replete medium, Nostoc sp. PCC 7120 cells were resilient even to prolonged P starvation when its growth was supported by mobilization of cellular reserves of P (polyphosphate) and glycogen on the background of accumulation of nitrogen and carbon reserves (mainly cyanophycin). The P-starving cells quickly recovered upon re-feeding with inorganic phosphate (Pi). Under diazotrophic conditions, P starvation essentially diminished the fixation of dinitrogen. As a result, most of the vegetative cells comprising the trichomes of the cyanobacterium died and decomposed while other cells retained their structural integrity but did not divide. In turn, the latter fell into two categories: some of them showed signs of nutrient starvation; while the other became dormant but did not display the signs of starvation. They resembled neither akinete nor chlorotic cells but were similar to arthrospores lacking a thickened sheath. Re-feeding with Pi triggered a quick resuscitation of the dormant vegetative cells manifested by mobilization of their internal reserves, resumption of the cell growth and division. These processes took place faster than the formation of heterocytes with well-developed envelope (thus, nitrogenase activity recovered by the 7th day after re-feeding of the cells with Pi). The results provide a deeper insight into the mechanisms of stress tolerance in Nostoc sp. PCC 7120 and modulation of the cyanobacterial productivity in natural ecosystems and artificial cultivation facilities by nitrogen and P availability.

Phosphorus (P) is an essential macronutrient central to the exchange and storage of energy and information in the cell. Due to its limited bioavailability, P often becomes a limiting nutrient in aquatic and terrestrial ecosystems hence the studies of responses to stress caused by P starvation cyanobacteria, the primary producers, are of considerable interest. Indeed, the availability of P is among main factors limiting diazotrophy in cyanobacteria. To gain a deeper understanding of the effect of P starvation on cyanobacteria in non-diazotrophic conditions, we studied a model system of two near—isogenic strains, Nostoc sp. PCC 7120 and Nostoc sp. PCC 7118 differ in their ability to form heterocysts. Specifically, we investigated the differences in the responses of these strains to P starvation by comparing their growth kinetics, photosynthetic pigment content, ultrastructural rearrangements of vegetative cells, and the expression profile of key genes of phosphorus metabolism. The tolerance of PCC 7120 to P starvation was higher than that of PCC 7118, which manifested itself in a higher growth rate, less profound ultrastructural changes (in particular, phycobilisomes as well as polyphosphate reserves were retained in the cells of PCC 7120). At the same time, the accumulation of cyanophycin, a depot of nitrogen and energy, increased several-fold in the cells of both strains during P starvation, but this increase was larger in PCC 7118 cells. Assumably, the increased resilience of the PCC 7120 to P starvation stems from its higher ability to accumulate intracellular reserves of P in the form of polyphosphates. Our findings suggest that the phenotypic differences between the strains Nostoc sp. PCC 7118 and Nostoc sp. PCC 7120 are not limited to the different ability to form heterocysts. A deeper understanding of the drivers of stress response phenotypic diversity in near-isogenic strains would require a comparative analysis of their whole-genome sequences.

Microalgae are naturally adapted to the fluctuating availability of phosphorus (P) to opportunistically uptake large amounts of inorganic phosphate (P-i) and safely store it in the cell as polyphosphate. Hence, many microalgal species are remarkably resilient to high concentrations of external P-i. Here, we report on an exception from this pattern comprised by a failure of the high P-i-resilience in strain Micractinium simplicissimum IPPAS C-2056 normally coping with very high P-i concentrations. This phenomenon occurred after the abrupt re-supplementation of P-i to the M. simplicissimum culture pre-starved of P. This was the case even if P-i was re-supplemented in a concentration far below the level toxic to the P-sufficient culture. We hypothesize that this effect can be mediated by a rapid formation of the potentially toxic short-chain polyphosphate following the mass influx of P-i into the P-starved cell. A possible reason for this is that the preceding P starvation impairs the capacity of the cell to convert the newly absorbed P-i into a "safe" storage form of long-chain polyphosphate. We believe that the findings of this study can help to avoid sudden culture crashes, and they are also of potential significance for the development of algae-based technologies for the efficient bioremoval of P from P-rich waste streams.

Due to their valuable properties, nanoparticles are in demand in industry, medicine, and ecology. Traditional physical-chemical synthesis methods of nanoparticles are energy-consuming and/or are carried out with the participation of toxic substances that limit their further use, e.g., in medicine. In this regard, attention is drawn to the “green” synthesis of nanoparticles including with the participation of microalgae cells. The ability of cells of the green microalgae Micractinium simplicissimum IPPAS C-2056 to form ultra-small Fe (1–4 nm) and P (3–9 nm) nanoparticles, Mn (10–60 nm) nanoparticles as well as mixed (P–Fe, P‒Mn, Fe–Mn, Fe–P–Ca–Mg) nanoparticles, which vary in composition, localized in the intercellular matrix, on the surface, and in the cell wall is shown by transmission electron microscopy combined with energy-dispersive X-ray spectroscopy. The obtained data evidence a high potential of M. simplicissimum IPPAS C2056 for the green synthesis of nanoparticles containing Mn, Fe, and P. The determinants of this ability and prospects for using microalgae to produce nanoparticles with valuable properties are discussed.

A novel chlorophyte algae strain with outstanding resilience to high inorganic phosphate (Pi) concentrations in the medium was isolated from a phosphorus-polluted site near a rock phosphate mine. According to the morphological, ultrastructural and genetic criteria the strain was assigned to the species Micractinium simplicissimum H.Chae, H.-G. Choi & J.H.Kim. This strain retained cell viability and growth capacity in the presence of Pi concentrations up to 14 g L–1. The uptake of Pi by the cells was moderate (equal to ca. 0.7% increase in cell dry weight P percentage) regardless of the amount of the exogenic Pi added to the culture. At the same time, approximately a half of the Pi removed by the M. simplicissimum from the culture was reversely adsorbed by the cell surface and/or the intercellular matrix and cell debris. The ultrastructural studies indicated the metabolically active status of the cells together with the presence of phosphorus-rich (likely, polyphosphate) inclusions outside and inside the cells (mainly in vacuoles). We hypothesized that the Pi resilience of the studied strain stems from its high Pi adsorption capacity together with its ability to throttle the Pi influx into the cell preventing the rapid buildup of intracellular Pi and potentially toxic short-chain polyphosphate.

Green microalgae Lobosphaera sp. IPPAS С-2047 and Micractinium simplicissimum IPPAS С-2056 were examined for the first time for cell tolerance to elevated concentrations of manganese applied in the form of MnCl2. Analyses of cell photosynthetic activity by chlorophyll fluorometry and the dynamic patterns of absorbance changes of cell suspensions at the peak of chlorophyll absorption revealed differential tolerance of two microalgae species to manganese. The acute toxicity assayed in 4-day treatments became apparent at manganese concentrations equal to or higher than 1 g/L for M. simplicissimum cells and at concentrations above 10 g/L for Lobosphaera sp. Transmission electron microscopy and energy-dispersive X-ray spectroscopy were used to study the subcellular distribution of manganese in microalgal cells under elevated nontoxic concentrations of manganese in the medium. The results with Lobosphaera sp. established the lack of individual manganese-containing inclusions on cell surfaces and in the cell interior; the intracellular distribution of manganese was dispersed with elevated accumulation of this element in the region of thylakoids and plastoglobules. The occurrence of manganese and phosphorus in plastoglobules was found for the first time. Apparently, these compartments become accessible for accumulation of Mn and P upon the translocation of thylakoid components during stress-induced disassembling of their structure. The cells of M. simplicissimum were able to oxidize the exoplasmic Mn2+ with the formation of manganese nanoparticles in the intercellular matrix as well as on the cell surface and within the cell walls. In addition, the manganese permeating into the cells was shown to compartmentalize in vacuoles and bind to the polyphosphate granules.

The immobilization of cells of a mixed culture of the microalgae (MA) Micractinium sp. NAMSU A-19 and cyanobacteria (CB) Synechococcus sp. 1Dp66E-1 on a chitosan-based polymer was studied. A polycationic sorbent based on a natural chitosan polymer with a molecular weight of 600 kDa was obtained from the crosslinking of chitosan with glutaraldehyde via cryopolymerization. It has a high affinity for the surface structures of oxygenic phototrophic microorganisms (OPMs) and enables strong cellular attachment to the sorbent surface. The study of the kinetics and evaluation of the effectiveness of mixed-culture immobilization showed a high sorption capacity of the chitosan sorbent. During the first hour of cultivation, the immobilization efficiency was on average 40–52%, and, almost all cells were immobilized after 48 h. The highly porous, nontoxic, and biodegradable sorbent provided steady cellular attachment for 7 days of cultivation and did not affect the growth of immobilized mixed culture, either on the surface or in the inner layers of the polymer. The study of mixed-culture immobilization via scanning electron microscopy showed that CB and MA cells are tightly attached to the surface of the chitosan sorbent. This is followed by the formation of strands of the extracellular polymer matrix and a biofilm consisting of cells of the mixed culture of MA and CB and heterotrophic bacteria associated with Micractinium sp. Immobilization on a chitosan sorbent contributes to an increase in the bioremoval of nitrates and phosphates by the tested mixed culture.

Biotechnology of microalgae holds promise for sustainable using of phosphorus, a finite non-renewable resource. Responses of the green microalga Lobosphaera sp. IPPAS C-2047 to elevated inorganic phosphate (Pi) concentrations were studied. Polyphosphate (PolyP) accumulation and ultrastructural rearrangements were followed in Lobosphaera using light and electron microscopy and linked to the responses of the photosynthetic apparatus probed with chlorophyll fluorescence. High tolerance of Lobosphaera to ≤ 50 g L–1 Pi was accompanied by a retention of photosynthetic activity and specific induction of non-photochemical quenching (NPQ up to 4; Fv/Fm around 0.7). Acclimation of the Lobosphaera to the high Pi was accompanied by expansion of the thylakoid lumen and accumulation of the carbon-rich compounds. The toxic effect of the extremely high (100 g L–1) Pi inhibited the growth by ca. 60

A range of microalgal cultures was tested to perform phosphorus luxury uptake and form the artificial microbial communities – the biofloccules of phosphorus accumulators.

MAIN CONCLUSION:Haematococcus lacustris inhabits supralittoral rock ponds and forms, under natural conditions, biofilms including layered cyanobacterial and fermentative microbial mats. Dry mats, formed under extremely stressful conditions, contained only haematocysts. Under favorable growth conditions, modeled for dry biofilms in vitro, microalgal free-living stages were detected. Haematococcus lacustris is the microalga known for its high potential to survive under a wide range of unfavorable conditions, particularly in the supralittoral temporal rock ponds of the White Sea. Previously, we described microbial communities containing H. lacustris in this region. In many cases, they were organized into systems exhibiting complex three-dimensional structure similar to that of natural biofilms. In this study, for the first time, we clarify structural description and provide microscopic evidence that these communities of H. lacustris and bacteria are assembled into the true biofilms. There are (1) simple single layer biofilms on the surface of rocks and macrophytic algae, (2) floccules (or flocs) not attached to a surface, (3) as well as stratified (layered) biofilms, wet, and dehydrated in nature. Being involved into primary organic production, H. lacustris and cyanobacteria are located exclusively in the upper layers of stratified biofilms, where they are capable to absorb sufficient for photosynthesis amount of light. The presence of acidic polysaccharides in the extracellular matrix revealed by specific staining with ruthenium red in the H. lacustris-containing microbial communities is a biochemical evidence of biofilm formation. Meanwhile, the presence of bacterial L-form is an ultrastructural confirmation of that fact. Under favorable conditions, modeled in vitro, H. lacustris from the dry microbial mats moves to the free-living states represented by vegetative palmelloid cells and motile zoospores. Owing to the fact that inside biofilms cells of microorganisms exist under stable conditions, we consider the biofilm formation as an additional mechanism that contributes to the survival of H. lacustris in the supralittoral zone of the White Sea.

Green microalgae are able to sustain hydrogen evolution in the light under nutrient deprivation conditions. In the current study, the role of hydrogen photoproduction in the acclimation of Chlamydomonas reinhardtii cells to nutrient deficiency was investigated. In this regard, such physiological characteristics of algal cells as starch and adenosine triphosphate content, fatty acid composition, fermentation products, photosynthetic activity, cell mortality, culture recovery, and cell ultrastructure were compared between the hydEF-1 mutant lacking hydrogenase activity and its parent strain CC-425 under sulfur deficiency. The results showed an increased number of non-viable cells, altered cell ultrastructure, and modified lipid, carbohydrate, and energy metabolism in the mutant cells. We suggest that active hydrogenase provides benefits for algae acclimation to the anaerobic phase of nutrient deprivation by regulating intracellular redox and pH balance and improving performance of anaerobic metabolism.

The internal surface of the animal gastrointestinal tract is covered by microbial biofilms. They play an important role in the development and functioning of the host organism and protect it against pathogens. Microbial communities of gastrointestinal biofilms are less elucidated than luminal microbiota. Therefore, the studies of biofilm formation by gastrointestinal microorganisms are a topical issue. For the first time, we report the formation of a biofilm in vitro by the strains of bioluminescent bacteria isolated from the intestines of marine fish. These bacteria exhibit co-aggregation and tend to attach to solid surfaces. The attachment of cells is accompanied by appearance of the pili. Then, we observed the formation of microcolonies and the production of extracellular polymer substances (EPSs) connecting bacterial cells into an integrated system. The presence of acidic polysaccharides is shown in the EPS when using the ruthenium red staining. Acidic polysaccharides in this matrix is a biochemical evidence of microbial biofilms. On the fibers of the polymer matrix, these bacteria form the "mushroom body"-type structures. Matured biofilms exhibit a specific three-dimensional architecture with pores and channels formed by cells and EPS. We also demonstrated the formation of a biofilm by binary culture of the luminous enterobacterium Kosakonia cowanii and a Gram-positive Macrococcus sp. The data obtained help to understand the role of these bacteria in the intestines of fish. They lead to a new study in the field of investigation of the intestinal microbiome of fish.

To cope with fluctuating phosphorus (P) availability, cyanobacteria developed diverse acclimations, including luxury P uptake (LPU)—taking up P in excess of the current metabolic demand. LPU is underexplored, despite its importance for nutrient-driven rearrangements in aquatic ecosystems. We studied the LPU after the refeeding of P-deprived cyanobacterium Nostoc sp. PCC 7118 with inorganic phosphate (Pi), including the kinetics of Pi uptake, turnover of polyphosphate, cell ultrastructure, and gene expression. The P-deprived cells deployed acclimations to P shortage (reduction of photosynthetic apparatus and mobilization of cell P reserves). The P-starved cells capable of LPU exhibited a biphasic kinetic of the Pi uptake and polyphosphate formation. The first (fast) phase (1–2 h after Pi refeeding) occurred independently of light and temperature. It was accompanied by a transient accumulation of polyphosphate, still upregulated genes encoding high-affinity Pi transporters, and an ATP-dependent polyphosphate kinase. During the second (slow) phase, recovery from P starvation was accompanied by the downregulation of these genes. Our study revealed no specific acclimation to ample P conditions in Nostoc sp. PCC 7118. We conclude that the observed LPU phenomenon does not likely result from the activation of a mechanism specific for ample P conditions. On the contrary, it stems from slow disengagement of the low-P responses after the abrupt transition from low-P to ample P conditions.

Bioluminescence is a spectacular feature of some prokaryotes. In the present work, we address the distribution of bioluminescence among bacteria isolated from the White Sea finfishes. Luminous bacteria are widely distributed throughout the World Ocean. Many strains have been isolated and described for tropical latitudes, while Nordic seas still remain quite a white spot in studying bioluminescence of bacteria. We describe the strains related to the two main genera of luminous bacteria, Photobacterium and Aliivibrio, as well as Shewanella and Vibrio. They are related to families Vibrionaceae and Shewanellaceae of the Gammaproteobacteria class. Here, we at the first time, report the bioluminescence of the Enterobacteriaceae Kosakonia cowanii. Moreover, we applied the polyphasic approach to identify and describe the isolated microorganisms. The data on sequencing, diversity of cell fine structure, and light emission spectra at room temperature on the solid medium are discussed. The bacteria are characterized by features in their light emission spectra. It may reflect possible molecular mechanisms of bioluminescence as well as features of bacterial composition. The obtained data expands the existing body of knowledge about the bioluminescence spread among the bacteria of Nordic latitudes and provides complex information that is crucial for their precise identification.

Algal cells are highly complex optical systems that can dynamically change their structure. Consequently, absorption and scattering properties of algae change, while the cells are acclimating to different light conditions or during growth and division in a cell cycle. This may be particularly important in algal species that can grow rapidly under very high-light such as Chlorella vulgaris IPPAS C-1 that is studied here. From cell transmittance measured conventionally and using integrating sphere, we evaluated absorption and scattering coefficients and cross sections per cell dry weight and chlorophyll content. This was done for asynchronous cell culture grown in low-light (LL; 220 μmol (photons) m−2 s−1) or high-light (HL; 1760 μmol (photons) m−2 s−1) light, as well as during cell cycle of synchronous culture grown in HL. During the cell cycle, we also determined cell ultrastructural organization by transmission electron microscopy, and correlated its parameters with absorption and scattering cross sections per cell dry weight. We found that the IPPAS C-1 cells of asynchronous culture scatter light more than other cells, however, internal organization of the cells that is decisive for scattering is less sensitive to HL and LL treatment than the cell pigment content that controls absorption. The light scattering and absorption were dynamically changed during cell cycle of synchronous cells grown in the HL. Changes in ratio of chloroplast to protoplast area, reflecting amount of scattering chloroplast membrane (outer, inner) interfaces, best correlated with changes in light scattering. We suggest that the increased light scattering by the HL-acclimated IPPAS C-1 cells might be responsible for increased HL resilience reported in the literature. Biotechnological aspect of this study is that the scattering and absorption properties of phytoplankton cells ought to be calibrated for each particular growth phase or irradiance to which the cells are acclimated.

Mitochondria-targeted antioxidants (also known as 'Skulachev Ions' electrophoretically accumulated by mitochondria) exert anti-ageing and ROS-protecting effects well documented in animal and human cells. However, their effects on chloroplast in photosynthetic cells and corresponding mechanisms are scarcely known. For the first time, we describe a dramatic quenching effect of (10-(6-plastoquinonyl)decyl triphenylphosphonium (SkQ1) on chlorophyll fluorescence, apparently mediated by redox interaction of SkQ1 with Mn cluster in Photosystem II (PSII) of chlorophyte microalga Chlorella vulgaris and disabling the oxygen-evolving complex (OEC). Microalgal cells displayed a vigorous uptake of SkQ1 which internal concentration built up to a very high level. Using optical and EPR spectroscopy, as well as electron donors and in silico molecular simulation techniques, we found that SkQ1 molecule can interact with Mn atoms of the OEC in PSII. This stops water splitting giving rise to potent quencher(s), e.g. oxidized reaction centre of PSII. Other components of the photosynthetic apparatus proved to be mostly intact. This effect of the Skulachev ions might help to develop in vivo models of photosynthetic cells with impaired OEC function but essentially intact otherwise. The observed phenomenon suggests that SkQ1 can be applied to study stress-induced damages to OEC in photosynthetic organisms.

In oxygenic phototrophs including unicellular algae, acclimation to and damage by diverse environmental stresses induce profound changes in the ultrastructural organization of the cell. These alterations reflect acclimation of the photosynthetic apparatus to unfavorable conditions (mainly reduction of the chloroplast and its membranal system) and rewiring of the photo-fixed carbon fluxes in the cell. These changes, eventually pursuing mitigation of the photooxidative damage risk, are manifested by the formation of diverse carbon-rich inclusions. Although the physiological and molecular basis of these processes are well understood, the ultrastructural manifestations of the stress responses are often fragmented and frequently controversial. This minireview attempts to generalize on the ultrastructural patterns accompanying stresses in the photosynthetic cell, involving the concerted rearrangements of its assimilatory and storage compartments. The changes characteristic of normal functioning and emergency reduction of the chloroplast thylakoids under harsh stress are also addressed. Special attention is paid to the manifestations of the engagement of photoprotection via active (energy-dependent non-photochemical quenching) and passive mechanisms (e.g. optical shielding by secondary carotenoids). We also underline the potentially important role of autophagy-like processes and provide a more integral view of ultrastructural rearrangements under stress.

We established a new simple approach to study phosphorus (P) and nitrogen (N) reserves at subcellular level potentially applicable to various types of cells capable of accumulating P- and/or N-rich inclusions. Here, we report on using this approach for locating and assessing the abundance of the P and N reserves in microalgal and cyanobacterial cells. The approach includes separation of the signal from P- or N-rich structures from noise on the energy-filtered transmission electron microscopy (EFTEM) P- or N-maps. The separation includes (i) relative entropy estimation for each pixel of the map, (ii) binary thresholding of the map, and (iii) segmenting the image to assess the inclusion relative area and localization in the cell section. The separation is based on comparing the a posteriori probability that a pixel of the map contains information about the sample vs. Gaussian a priori probability that the pixel contains noise. The difference is expressed as relative entropy value for the pixel; positive values are characteristic of the pixels containing the payload information about the sample. This is the first known method for quantification and locating at a subcellular level P-rich and N-rich inclusions including tiny (< 180 nm) structures. We demonstrated the applicability of the proposed method both to the cells of eukaryotic green microalgae and cyanobacteria. Using the new method, we elucidated the heterogeneity of the studied cells in accumulation of P and N reserves across different species. The proposed approach will be handy for any cytological and microbiological study requiring a comparative assessment of subcellular distribution of cyanophycin, polyphosphates or other type of P- or N-rich inclusions. An added value is the potential of this approach for automation of the data processing and evaluation enabling an unprecedented increase of the EFTEM analysis throughput.