Inna Khozin-Goldberg

This study uncovers the critical role of dietary n-3 polyunsaturated fatty acids (PUFAs) in the development of dormant embryos, known as resting eggs (REs), in the aquatic rotifer Brachionus plicatilis. REs contained PUFAs, and the maternal diet markedly influenced their levels. In this respect, supplementing PUFAs-deficient yeast with eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) in the diet increased RE production. A combination of EPA and DHA yielded the highest number of produced REs and reduced the percentage of damaged REs, without altering hatching rates. Proteome profiling investigated the impact of EPA and DHA on protein expression patterns. It identified 283 differentially expressed proteins out of 1954 and indicated distinct EPA and DHA functions. Differentially expressed proteins were linked to multiple functional pathways, suggesting nuclear receptor regulation. In vertebrates, EPA, DHA, and α-linolenic acid (ALA) interact with retinoid and peroxisome proliferator-activated receptors. These receptors emerged as promising regulators in PUFAs-treated rotifers. Consequently, the impact of ALA was also assessed. Remarkably, ALA enhanced reproductive rates, increased RE production, reduced the proportion of damaged REs, and improved hatching in a dose-dependent manner, even in the absence of EPA and DHA. These findings emphasize the essential roles of PUFAs in rotifer RE formation, a previously unexplored topic. The study stresses the need to investigate the roles of PUFAs during dormancy. This is important because global warming may reduce the availability of PUFAs, decrease the quantity of REs in egg banks, and affect the long-term viability of aquatic non-vertebrate populations.

Fish production in recirculating aquaculture systems (RAS) continues to increase. Fish assimilate 20-30% of the applied feed, the rest is excreted mainly as ammonia and solids. In RAS, toxic ammonia undergoes nitrification to the much less toxic nitrate, which is removed through water exchange or through denitrification, and solids are separated and discharged. The aim of this study was to develop and validate a novel water treatment for RAS, where fish excretions are utilized for microalgal biomass production. A near zero-discharge microalgae-based RAS (mic-RAS) was established, integrating wastewater treatment with microalgal production, within a fully closed loop. A photobioreactor inoculated with the eustigmatophyte microalga BZ-1 was operated as the sole treatment unit, directly assimilating dissolved and particulate fish waste.Stable continuous operation was demonstrated at Cyprinus rubrofuscus (koi fish) stocking densities of 10–20 g L⁻¹ and a microalgal biomass concentration of 1 g L⁻¹ under illumination of 400 μmol photons m⁻² s⁻¹. Under these conditions, total ammonia nitrogen (TAN), nitrite, and nitrate remained below toxicity thresholds. Hydraulic stability was maintained through optimized air-assisted membrane backwash. In parallel with wastewater treatment, the mic-RAS generated valuable microalgal biomass rich in the long-chain polyunsaturated fatty acids eicosapentaenoic acid (EPA) and arachidonic acid (AA), demonstrating direct coupling with nitrogen removal. By embedding waste assimilation, hydraulic control, and biomass recovery, the mic-RAS shifts nutrient management from disposal toward resource retention and valorization. These results provide quantitative design constraints and demonstrate a scalable pathway toward more resource-efficient, circular, and environmentally sustainable aquaculture systems.

Phosphorus (P) is an important and irreplaceable macronutrient. It is central to energy and information storage and exchange in living cells. P is an element with a “broken geochemical cycle” since it lacks abundant volatile compounds capable of closing the P cycle. P fertilizers are critical for global food security, but the reserves of minable P are scarce and non-evenly distributed between countries of the world. Accordingly, the risks of global crisis due to limited access to P reserves are expected to be graver than those entailed by competition for fossil hydrocarbons. Paradoxically, despite the scarcity and value of P reserves, its usage is extremely inefficient: the current waste rate reaches 80% giving rise to a plethora of unwanted consequences such as eutrophication leading to harmful algal blooms. Microalgal biotechnology is a promising solution to tackle this challenge. The proposed review briefly presents the relevant aspects of microalgal P metabolism such as cell P reserve composition and turnover, and the regulation of P uptake kinetics for maximization of P uptake efficiency with a focus on novel knowledge. The multifaceted role of polyPhosphates, the largest cell depot for P, is discussed with emphasis on the P toxicity mediated by short-chain polyPhosphates. Opportunities and hurdles of P bioremoval via P uptake from waste streams with microalgal cultures, either suspended or immobilized, are discussed. Possible avenues of P-rich microalgal biomass such as biofertilizer production or extraction of valuable polyPhosphates and other bioproducts are considered. The review concludes with a comprehensive assessment of the current potential of microalgal biotechnology for ensuring the sustainable usage of phosphorus.

Phosphorus (P) is an important and irreplaceable macronutrient. It is central to energy and information storage and exchange in living cell. P is an element with “broken geochemical cycle” since it lacks abundant volatile compounds capable of closing the P cycle. P fertilizers are critical for global food security, but the reserves of minable P are scarce and non-evenly distributed between countries of the world. Accordingly, the risks of global crisis due to limited access to P reserves is expected to be graver than those entailed by competition for fossil hydrocarbons. Paradoxically, despite the scarcity and value of P reserves, its usage is extremely inefficient: current waste rate reaches 80% giving rise to a plethora of unwanted consequences such as eutrophication leading to harmful algal blooms. Microalgal biotechnology comprise a promising solution to tackle this challenge. The proposed review briefly presents the relevant aspects of microalgal P metabolism such as cell P reserve composition and turnover, and the regulation of P uptake kinetics for maximization of P uptake efficiency with the focus on novel knowledge. Multifaceted role of polyPhosphates, the largest cell depot for P, is discussed with emphasis on the P toxicity mediated by short-chain polyPhosphates. Opportunities and hurdles of P bioremoval via P uptake from waste streams with microalgal cultures, either suspended or immobilized, is discussed along with the role of bacterial components of microalgal-bacterial consortia in these processes. Possible avenues of P-rich microalgal biomass such as biofertilizer production or extraction of valuable polyPhosphates and other bioproducts are considered. The review concludes with a comprehensive assessment of the current potential of microalgal biotechnology for ensuring the sustainable usage of phosphorus.

Phaeodactylum tricornutum is a model oleaginous pennate diatom, widely investigated for the accumulation of triacylglycerols (TAG) in lipid droplets during nitrogen (N) starvation. However, lipid droplet breakdown, TAG catabolism, and remobilization upon N replenishment during growth restoration are less studied. Serine hydrolases (SH) constitute a diverse family encompassing proteases, amidases, esterases, and lipases. In this report, we adopted a chemoproteomic approach called Activity-Based Protein Profiling (ABPP) to explore the repertoire of active serine hydrolases to elucidate the mechanisms of lipid metabolism in P. tricornutum (strain Pt4). A superfamily-wide profile of serine hydrolases revealed a differentially active proteome (activome) during N starvation and after nutrient replenishment. We report 30 active serine hydrolases, which were broadly categorized into metabolic serine hydrolases and serine proteases. Lipases appeared to be the major metabolic linchpins prevalent during lipid remobilization. Global transcriptomics analysis provided a complementary insight into the gene expression level of the detected serine hydrolases. It revealed putative phospholipases as central players in membrane lipid turnover and remodeling involved in cellular lipid homeostasis and TAG accumulation. TAG remobilization and lipid droplet breakdown were impaired in the presence of phenyl mercuric acetate (PMA), whose activity as an SH inhibitor was validated by competitive ABPP. Lipid species profiling corroborated the impairment in TAG degradation and the buildup of structural lipids in the presence of PMA after nutrient replenishment. Collectively, our functional proteome approach, coupled with the transcriptome and lipidome data, provides a comprehensive landscape of bona fide active serine hydrolases, including lipases in this model diatom.

To feed the rapidly increasing world population, food production will have to double while the amount of water and arable land remain unchanged. Therefore, we must transition to environmentally sustainable, but just as nutritious diets by 2050. This will require drastic reduction in animal-based foods and increasing plant-based foods. Plant- based diets are more sustainable and healthier, with but one exception, plants are low in cobalamin (vitamin B ₁₂ ). Only a few plant and fungi species contain cobalamin, and some of the cobalamin forms present in algae and mushrooms are not available to humans. Recently, the smallest known plant, the edible duckweed Wolffia globosa Mankai, was shown to contain high concentrations of bioavailable cobalamin. We hypothesized that the production of bioavailable cobalamin is not unique to W. globosa Mankai but is common to other duckweed species. We also hypothesized that cobalamin production depends on the conditions under which the duckweed is grown. To test our hypotheses, we cultivated nine duckweed species under mixotrophic and phototrophic conditions, then measured the concentration and identified the bioavailability of cobalamin using biological and analytical methods. Our results showed that all duckweed species tested produce bioavailable cobalamin while pseudocobalamin (the non-available form of cobalamin) was not detected. We also found that mixotrophic conditions enhanced duckweed growth but reduced cobalamin concentrations. Our results establish duckweed species as a novel source of plant-derived bioavailable cobalamin and highlight the effect of growth conditions on production. More research is needed to identify the bacteria responsible for the production of cobalamin in duckweed and to optimize the conditions for cobalamin production.

Diatoms (Bacillariophyceae) accumulate neutral storage lipids in lipid droplets during stress conditions, which can be rapidly degraded and recycled when optimal conditions resume. Since nutrient and light availability fluctuate in marine environments, storage lipid turnover is essential for diatom dominance of marine ecosystems. Diatoms have garnered attention for their potential to provide a sustainable source of omega-3 fatty acids. Several independent proteomic studies of lipid droplets isolated from the model oleaginous pennate diatom Phaeodactylum tricornutum have identified a previously uncharacterized protein with an acyl-CoA binding (ACB) domain, Phatrdraft_48778, here referred to as Phaeodactylum tricornutum acyl-CoA binding protein (PtACBP). We report the phenotypic effects of CRISPR-Cas9 targeted genome editing of PtACBP. ptacbp mutants were defective in lipid droplet and triacylglycerol degradation, as well as lipid and eicosapentaenoic acid synthesis, during recovery from nitrogen starvation. Transcription of genes responsible for peroxisomal β-oxidation, triacylglycerol lipolysis, and eicosapentaenoic acid synthesis was inhibited. A lipid binding assay using a synthetic ACB domain from PtACBP indicated preferential binding specificity towards certain polar lipids. PtACBP fused to eGFP displayed an endomembrane-like pattern, which surrounded the periphery of lipid droplets. PtACBP is likely responsible for intracellular acyl transport, affecting cell division, development, photosynthesis, and stress response. A deeper understanding of the molecular mechanisms governing storage lipid turnover will be crucial for developing diatoms and other microalgae as biotechnological cell factories.