Crypthecodinium cohnii is a marine heterotrophic dinoflagellate that can accumulate high amounts of omega-3 polyunsaturated fatty acids (PUFAs), and thus has the potential to replace conventional PUFAs production with eco-friendlier technology. So far, C. cohnii cultivation has been mainly carried out with the use of yeast extract (YE) as a nitrogen source. In the present study, alternative carbon and nitrogen sources were studied: the extraction ethanol (EE), remaining after lipid extraction, as a carbon source, and dinoflagellate extract (DE) from recycled algae biomass C. cohnii as a source of carbon, nitrogen, and vitamins. In mediums with glucose and DE, the highest specific biomass growth rate reached a maximum of 1.012 h⁻¹, while the biomass yield from substrate reached 0.601 g·g⁻¹. EE as the carbon source, in comparison to pure ethanol, showed good results in terms of stimulating the biomass growth rate (an 18.5% increase in specific biomass growth rate was observed). DE supplement to the EE-based mediums promoted both the biomass growth (the specific growth rate reached 0.701 h⁻¹) and yield from the substrate (0.234 g·g⁻¹). The FTIR spectroscopy data showed that mediums supplemented with EE or DE promoted the accumulation of PUFAs/docosahexaenoic acid (DHA), when compared to mediums containing glucose and commercial YE.

Docosahexaenoic acid (DHA) is one of the most important long-chain polyunsaturated fatty acids (LC-PUFAs), with numerous health benefits. Crypthecodinium cohnii, a marine heterotrophic dinoflagellate, is successfully used for the industrial production of DHA because it can accumulate DHA at high concentrations within the cells. Glycerol is an interesting renewable substrate for DHA production since it is a by-product of biodiesel production and other industries, and is globally generated in large quantities. The DHA production potential from glycerol, ethanol and glucose is compared by combining fermentation experiments with the pathway-scale kinetic modeling and constraint-based stoichiometric modeling of C. cohnii metabolism. Glycerol has the slowest biomass growth rate among the tested substrates. This is partially compensated by the highest PUFAs fraction, where DHA is dominant. Mathematical modeling reveals that glycerol has the best experimentally observed carbon transformation rate into biomass, reaching the closest values to the theoretical upper limit. In addition to our observations, the published experimental evidence indicates that crude glycerol is readily consumed by C. cohnii, making glycerol an attractive substrate for DHA production.

Purine auxotrophy is an abundant trait among eukaryotic parasites and a typical marker for many budding yeast strains. Supplementation with an additional purine source (such as adenine) is necessary to cultivate these strains. If not supplied in adequate amounts, purine starvation sets in. We explored purine starvation effects in a model organism, a budding yeast Saccharomyces cerevisiae ade8 knockout, at the level of cellular morphology, central carbon metabolism, and global transcriptome. We observed that purine-starved cells stopped their cycle in G1/G0 state and accumulated trehalose, and the intracellular concentration of AXP decreased, but adenylate charge remained stable. Cells became tolerant to severe environmental stresses. Intracellular RNA concentration decreased, and massive downregulation of ribosomal biosynthesis genes occurred. We proved that the expression of new proteins during purine starvation is critical for cells to attain stress tolerance phenotype Msn2/4p targets are upregulated in purine-starved cells when compared to cells cultivated in purine-rich media. The overall transcriptomic response to purine starvation resembles that of stationary phase cells. Our results demonstrate that the induction of a strong stress resistance phenotype in budding yeast can be caused not only by natural starvation, but also starvation for metabolic intermediates, such as purines.

The incidence of autoimmunity is increasing, to ensure timely and comprehensive treatment, there must be a diagnostic method or markers that would be available to the general public. Fourier-transform infrared spectroscopy (FTIR) is a relatively inexpensive and accurate method for determining metabolic fingerprint. The metabolism, molecular composition and function of blood cells vary according to individual physiological and pathological conditions. Thus, by obtaining autoimmune disease-specific metabolic fingerprint markers in peripheral blood mononuclear cells (PBMC) and subsequently using machine learning algorithms, it might be possible to create a tool that will allow the diagnosis of autoimmune diseases. In this preliminary study, it was found that the peak shift at 1545 cm⁻¹ could be considered specific for autoimmune disease type 1 diabetes (T1D), while the shifts at 1070 and 1417 cm⁻¹ could be more attributed to the autoimmune condition per se. The prediction of T1D, despite the small number of participants in the study, showed an inverse AUC = 0.33 ± 0.096, n = 15, indicating a stable trend in the prediction of T1D based on FTIR metabolic fingerprint data in the PBMC.

Understanding the energy metabolism and its regulation is one of the clues to metabolic engineering of stress-resistant lignocellulose-converting microbial strains, also including the promising ethanologen Zymomonas mobilis. Z. mobilis is an obligately fermentative, facultatively anaerobic bacterium, carrying an active respiratory chain with low energy-coupling efficiency. Its respiration does not supply energy to aerobically growing cultures on sugary media, yet oxidative phosphorylation has been demonstrated in non-growing cells with ethanol. Here, we show, for the first time, that in respiring, non-growing Z. mobilis cells receiving regular small amounts of ethanol, oxidative phosphorylation significantly contributes to the maintenance of their viability. No improvement of viability is seen in the NADH dehydrogenase (ndh)-deficient respiratory mutant, which is unable to oxidize ethanol. The ethanol effect is also hampered by the protonophoric uncoupler CCCP, or the inhibitor of ATP synthase, DCCD. At higher concentrations (6% v/v), ethanol causes stress that slows down culture growth. By monitoring the activity of several respiratory gene promoters under ethanol stress with the green fluorescent protein reporter system, we demonstrate downregulation of these promoters, in particular the ndh promoter. We speculate that the decrease in respiratory chain activity in response to stress conditions mitigates the production of reactive oxygen species.