Skeletonema marinoi ecotypes show specific habitat-related responses to fluctuating light supporting high potential for growth under photobioreactor light regime.

Diatoms are a diverse group of phytoplankton usually dominating areas characterized by rapidly shifting light conditions. Because of their high growth rates and interesting biochemical profile, their biomass is considered for various commercial applications. This study aimed at identifying strains with superior growth in a photobioreactor (PBR) by screening the natural intraspecific diversity of ecotypes isolated from different habitats. We investigated the effect of PBR light fluctuating on a millisecond scale (FL, simulating the light in a PBR) on 19 ecotypes of the diatom Skeletonema marinoi isolated from the North Sea-Baltic Sea area. We compare growth, pigment ratios, phylogeny, photo-physiological variables and photoacclimation strategies between all strains and perform qPCR and absorption spectra analysis on a subset of strains. Our results show that the ecotypes responded differently to FL, and have contrasting photo-physiological and photoprotective strategies. The strains from Kattegat performed better in FL, and shared common photoacclimation and photoprotection strategies that are the results of adaptation to the specific light climate of the Kattegat area. The strains that performed better with FL conditions had a high light (HL)-acclimated phenotype coupled with unique nonphotochemical quenching features. Based on their characteristics, three strains were identified as good candidates for growth in PBRs.

Zeaxanthin epoxidase 3 Knockout Mutants of the Model Diatom Phaeodactylum tricornutum Enable Commercial Production of the Bioactive Carotenoid Diatoxanthin.

Carotenoids are pigments that have a range of functions in human health. The carotenoid diatoxanthin is suggested to have antioxidant, anti-inflammatory and chemo-preventive properties. Diatoxanthin is only produced by a few groups of microalgae, where it functions in photoprotection. Its large-scale production in microalgae is currently not feasible. In fact, rapid conversion into the inactive pigment diadinoxanthin is triggered when cells are removed from a high-intensity light source, which is the case during large-scale harvesting of microalgae biomass. Zeaxanthin epoxidase (ZEP) 2 and/or ZEP3 have been suggested to be responsible for the back-conversion of high-light accumulated diatoxanthin to diadinoxanthin in low-light in diatoms. Using CRISPR/Cas9 gene editing technology, we knocked out the ZEP2 and ZEP3 genes in the marine diatom Phaeodactylum tricornutum to investigate their role in the diadinoxanthin-diatoxanthin cycle and determine if one of the mutant strains could function as a diatoxanthin production line. Light-shift experiments proved that ZEP3 encodes the enzyme converting diatoxanthin to diadinoxanthin in low light. Loss of ZEP3 caused the high-light-accumulated diatoxanthin to be stable for several hours after the cultures had been returned to low light, suggesting that zep3 mutant strains could be suitable as commercial production lines of diatoxanthin.

Loss of CpFTSY Reduces Photosynthetic Performance and Affects Insertion of PsaC of PSI in Diatoms.

The chloroplast signal recognition particle (CpSRP) receptor (CpFTSY) is a component of the CpSRP pathway that post-translationally targets light-harvesting complex proteins (LHCPs) to the thylakoid membranes in plants and green algae containing chloroplasts derived from primary endosymbiosis. In plants, CpFTSY also plays a major role in the co-translational incorporation of chloroplast-encoded subunits of photosynthetic complexes into the thylakoids. This role has not been demonstrated in green algae. So far, its function in organisms with chloroplasts derived from secondary endosymbiotic events has not been elucidated. Here, we report the generation and characterization of mutants lacking CpFTSY in the diatom Phaeodactylum tricornutum. We found that this protein is not involved in inserting LHCPs into thylakoid membranes, indicating that the post-translational part of the CpSRP pathway is not active in this group of microalgae. The lack of CpFTSY caused an increased level of photoprotection, low electron transport rates, inefficient repair of photosystem II (PSII), reduced growth, a strong decline in the PSI subunit PsaC and upregulation of proteins that might compensate for a non-functional co-translational CpSRP pathway during light stress conditions. The phenotype was highly similar to the one described for diatoms lacking another component of the co-translational CpSRP pathway, the CpSRP54 protein. However, in contrast to cpsrp54 mutants, only one thylakoid membrane protein, PetD of the Cytb6f complex, was downregulated in cpftsy. Our results point to a minor role for CpFTSY in the co-translational CpSRP pathway, suggesting that other mechanisms may partially compensate for the effect of a disrupted CpSRP pathway.

Functional studies of CpSRP54 in diatoms show that the mechanism of thylakoid protein insertion differs from that in plants and green algae.

The chloroplast signal recognition particle 54 kDa (CpSRP54) protein is a member of the CpSRP pathway known to target proteins to thylakoid membranes in plants and green algae. Loss of CpSRP54 in the marine diatom Phaeodactylum tricornutum lowers the accumulation of a selection of chloroplast-encoded subunits of photosynthetic complexes, indicating a role in the co-translational part of the CpSRP pathway. In contrast to plants and green algae, absence of CpSRP54 does not have a negative effect on the content of light-harvesting antenna complex proteins and pigments in P. tricornutum, indicating that the diatom CpSRP54 protein has not evolved to function in the post-translational part of the CpSRP pathway. Cpsrp54 KO mutants display altered photophysiological responses, with a stronger induction of photoprotective mechanisms and lower growth rates compared to wild type when exposed to increased light intensities. Nonetheless, their phenotype is relatively mild, thanks to the activation of mechanisms alleviating the loss of CpSRP54, involving upregulation of chaperones. We conclude that plants, green algae, and diatoms have evolved differences in the pathways for co-translational and post-translational insertion of proteins into the thylakoid membranes.

Loss of ALBINO3b Insertase Results in Truncated Light-Harvesting Antenna in Diatoms.

The family of chloroplast ALBINO3 (ALB3) proteins function in the insertion and assembly of thylakoid membrane protein complexes. Loss of ALB3b in the marine diatom Phaeodactylum tricornutum leads to a striking change of cell color from the normal brown to green. A 75% decrease of the main fucoxanthin-chlorophyll a/c-binding proteins was identified in the alb3b strains as the cause of changes in the spectral properties of the mutant cells. The alb3b lines exhibit a truncated light-harvesting antenna phenotype with reduced amounts of light-harvesting pigments and require a higher light intensity for saturation of photosynthesis. Accumulation of photoprotective pigments and light-harvesting complex stress-related proteins was not negatively affected in the mutant strains, but still the capacity for nonphotochemical quenching was lower compared with the wild type. In plants and green algae, ALB3 proteins interact with members of the chloroplast signal recognition particle pathway through a Lys-rich C-terminal domain. A novel conserved C-terminal domain was identified in diatoms and other stramenopiles, questioning if ALB3b proteins have the same interaction partners as their plant/green algae homologs.

Long-term dynamics, population structure and connectivity of the helmet jellyfish Periphylla periphylla in a Norwegian fjord and adjacent waters.

Mass occurrences of Periphylla periphylla in Norwegian fjords cause major concerns related to potential regime shifts that could affect ecosystem stability. 15 years of trawl data (2006-2015), complemented with comprehensive sampling in different areas and seasons (2018-2021) allowed new insights on the dynamics, structure and connectivity of P. periphylla populations within and beyond Trondheimsfjorden. Despite assumed population bursts, no clear trend on P. periphylla population size in Trondheimsfjorden were identified. Sampling frequency and population size suggest a local reproduction of P. periphylla, especially in the inner part of the fjord where young-of-the-year (YOY) individuals occur. Size variations occurred in relation to sampling month, thus pointing at seasonal patterns in growth and reproduction. No distinct population structure of P. periphylla populations within Trondheimsfjorden and over larger spatial scales (> 100 km) along the Norwegian coast was observed. Such poor geographic population structure provides evidence for a strong dispersal of P. periphylla, potentially triggered by frequent deep-water renewals of the fjords' basins that enable a high gene flow. Data on P. periphylla long-term dynamics, population structure and connectivity provide valuable information for ecosystem state assessments and enable the advancement of ecosystem management approaches, thus accounting for both stakeholder and ecosystem demands.

Nanocosm: a well plate photobioreactor for environmental and biotechnological studies.

Phytoplankton are key primary producers at the bottom of the aquatic food chain. They are a highly diverse group of organisms essential for the functioning of our ecosystems and because of their characteristics, their biomass is considered for various commercial applications. A full appreciation of their abundance, diversity and potential is only feasible by using systems that enable simultaneous testing of strains and/or variables in a fast and easy way. A major bottleneck is the lack of a cost-effective method with the capacity for complex experimental set-ups that enable fast and reproducible screening and analysis. In this study, we present nanocosm, a versatile LED-based micro-scale photobioreactor (PBR) that allows simultaneous testing of multiple variables such as temperature and light within the same plate. Every well can be independently controlled for intensity, temporal variation and light type (RGB, white, UV). We show that our systems guarantee homogeneous conditions because of controlled temperature and evaporation and adjustments for light crosstalk. By ensuring controlled environmental conditions the nanocosm is suitable for running factorial experimental designs where each well can be used as an independent micro-PBR. To validate culture performances, we assess well-to-well reproducibility and our results show minimal well-to-well variability for all the conditions tested. Possible modes of operation and application are discussed together with future development of the system.