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Closed photobioreactors reach temperatures that reduce microalgal production or even cause culture collapses. Cooling can maintain the temperature within tolerable boundaries, but cooling is energy‐intensive and expensive. Thermotolerant microalgal strains can reduce dependence on such cooling. In this study, adaptive laboratory evolution was performed for 390 days to further increase the maximal tolerable temperature for the already thermotolerant microalgae Picochlorum sp. (BPE23). The parental wild‐type strain of Picochlorum sp. (BPE23) exhibited a maximum mid‐day growth temperature of 47.5°C, whereas the isolated clones grew up to 49°C. At a lower temperature of 40°C, the growth rate and absorption cross‐sectional area were similar for the wild‐type strain and the evolved clones. Interestingly, the clones showed a 46% increase in cell volume compared to the wild‐type strain. The evolved clones with an expanded upper‐temperature boundary can be applied for broader temperature control of 1.5°C, without trade‐off effects at lower temperatures.

Photobioreactors heat up significantly during the day due to irradiation by sunlight. High temperatures affect cell physiology negatively, causing reduced growth and productivity. To elucidate the microalgal response to stressful supra-optimal temperature, we studied the physiology of Picochlorum sp. (BPE23) after increasing the growth temperature from 30 °C to 42 °C, whereas 38 °C is its optimal growth temperature. Cell growth, cell composition and mRNA expression patterns were regularly analyzed for 120 h after increasing the temperature. The supra-optimal temperature caused cell cycle arrest for 8 h, with concomitant changes in metabolic activity. Accumulation of fatty acids was observed during this period to store unspent energy which was otherwise used for growth. In addition, the microalgae changed their pigment and fatty acid composition. For example, palmitic acid (C16:0) content in the polar fatty acid fraction increased by 30%, hypothetically to reduce membrane fluidity to counteract the effect of increased temperature. After the relief of cell cycle arrest, the metabolic activity of Picochlorum sp. (BPE23) reduced significantly over time. A strong response in gene expression was observed directly after the increase in temperature, which was dampened in the remainder of the experiment. mRNA expression levels associated with pathways associated with genes acting in photosynthesis, carbon fixation, ribosome, citrate cycle, and biosynthesis of metabolites and amino acids were downregulated, whereas the proteasome, autophagy and endocytosis were upregulated.

Multiple models have been developed in the field to simulate growth and product accumulation of microalgal cultures. These models heavily depend on the accurate estimation of growth parameters. In this paper growth parameters are presented for three industrially relevant microalgae species: Nannochloropsis sp., Neochloris oleoabundans, and Picochlorum sp. (BPE23). Dedicated growth experiments were done in photobioreactors to determine the maximal biomass yield on light and maintenance rate, while oxygen evolution experiments were performed to estimate the maximal specific growth rate. Picochlorum sp. exhibited the highest specific growth rate of 4.98 ± 0.24 day −1 and the lowest specific maintenance rate of 0.079 day −1 , whereas N. oleoabundans showed the highest biomass yield on light of 1.78 g x •mol ph −1 . The measured growth parameters were used in a simple kinetic growth model for verification. When simulating growth under light conditions as found at Bonaire (12 °N, 68°W), Picochlorum sp. displayed the highest areal biomass productivity of 32.2 g.m −2 •day −1 and photosynthetic efficiency of 2.8%. The