ABSTRACT Although extraction of polysaccharides to convert reducing sugars (RS) from microalgae by acid or alkali pretreatments and enzymatic hydrolysis has been extensively studied, few reports exploring the use of high hydrostatic pressure processing (HHP) and ultrasonication (US) as emerging technologies for the extraction of sugars from microalgae biomass exist. Thus, the present study was conducted to determine the effects of mixotrophic growth and stress conditions (NaNO3 and CO2 concentration and light intensity) on RS and protein accumulation in the unicellular green alga Chlorella vulgaris in addition to optimization of the effectiveness of the sequential applications of HHP and US with dilute acid as well as simultaneous enzymatic saccharification on the production of RS from microalga cells. High light intensity, high CO2 concentration and limited nitrogen concentration promoted RS production. The maximum protein content (0.0683 mg g‒1) was achieved at 0.3 g l‒1 NaNO3 concentration, 7000 μmol photons m‒2 s‒1 and 6 l min‒1 CO2 concentration. The highest RS content of C. vulgaris after 48 h enzymatic saccharification (583.86 ± 13.23 mg g‒1) was obtained at 1% (w/w) acid concentration and 80% amplitude for 30 min with 79.4% RS yield. Combined US-assisted dilute acid pretreatment and enzymatic hydrolysis were also found to be more effective than HHP assisted dilute acid pretreatment and enzymatic saccharification. Therefore, microalgal biomass can be considered a suitable renewable feedstock used in fermentation.

ABSTRACT Although genome sequences of lichenized fungi are increasingly becoming available, genome sequences of microalgae involved in the lichen symbiosis are still scarce. For lichenized eukaryotic algae, genome sequencing has focused mostly on Trebouxia and Asterochloris, with little genomic data available for Stichococcus-like algae, such as Diplosphaera. The genus Diplosphaera is a common component of biological soil crusts, and often occurs associated with lichens of the family Verrucariaceae. It is characterized by cylindrical to spherical cells containing a plate-like chloroplast, and more specifically by a vegetative cell division that leads to the formation of typical two- to four-celled clusters. Here, we present a draft genome sequence for the algal partner of an Australian lichen specimen of Endocarpon pusillum. The genome was sequenced with Pac Bio long read and Illumina short read technologies, and transcriptome data were generated to inform the structural annotations. This algal strain is here identified as Diplosphaera chodatii based on nuSSU and ITS data. Compared with closely related lichenized and non-lichenized algae, the genome of D. chodatii stands out for its large size (85.6 Mb) and gene content (21,261 protein-encoding regions), as well as its high rate of duplicated genes (60% of the BUSCO genes are duplicated). These results suggest that whole genome duplication or large-scale segmental duplications may have occurred in the evolutionary history of this algal species. HIGHLIGHTS Little genome data are available for lichenized algae. We generated the first genome for a lichenized Diplosphaera chodatii. Results suggest a possible whole genome duplication in this species.

ABSTRACT Oceanic phytoplankton species are generally composed of many strains, with intra-species diversity consisting of genetic and phenotypic variability. Despite its importance in ecological and biotechnological contexts, this intra-species diversity and variation among strains has been little studied. We investigated the intra-species diversity of the microalga Tisochrysis lutea, a haptophyte of the Isochrysidales order. Inter-strain diversity of T. lutea was studied because of the economic importance of the species as a feed in aquaculture and for antioxidant metabolite production, particularly fucoxanthin and other carotenoids, which have health benefits. We analysed Tara Ocean datasets which revealed that T. lutea was present in the Pacific, Atlantic and Indian Oceans but not in the Arctic or Austral Oceans. We next made phenotypic and genotypic comparisons of 11 strains of T. lutea from worldwide algal collections. All strains were cultivated in the same controlled conditions for one week, and several phenotypic traits were measured, notably antioxidant content. In parallel, the genomes of each strain were sequenced, and genetic variants identified. At the genetic and phenotypic levels, the strains were distinct from each other and our analysis revealed natural trait variations of interest in relation to further exploitation in domestication programmes. A large number of genetic variations were identified among the strains, but no major differences in genome size were observed. Moreover, limited genetic structure was observed among these strains, which could be a consequence of the complex life history of species within the Isochrysidales. Our study provides new knowledge on the intra-species diversity that should be considered in future environmental studies and breeding programmes.

ABSTRACT The importance of diatoms in bioindication and palaeolimnology is significant. For these applications, and species identification in general, a sound understanding of morphological variability is essential. Some species of the centric diatom genera Cyclostephanos and Stephanodiscus can form long, chain-like colonies. In these genera colony formation is associated with the morphology of the spines, which hold the cells together. One objective of this study was to investigate other possible differences in the valve morphology of solitary and colonial cultures. In addition, all strains were sequenced to account for the possibility that solitary and colonial forms may be different species, using three different loci (LSU, rbcL, cox1). The results show that colony formation not only affects the spine morphology but also other morphological features of the valve. The current interpretation that C. invisitatus and C. delicatus are facultative colony formers is supported by the results of this study. The evidence furthermore suggests that C. incognitus is a morphotype of C. invisitatus. A similar morphological pattern leads to the hypothesis that S. hantzschii and S. binderanus could possibly be solitary and colonial forms of the same species, which is also in agreement with the molecular analysis. These findings have implications for the taxonomy of the group and its application and open up a way for future investigations of morphological changes due to colony formation in other groups and taxa.

ABSTRACT In this study we explore the introduction of the brown alga Dictyota acutiloba in the Mediterranean Sea and provide a substantive update on the geographic distribution of this species, which was long thought to be confined to the Pacific Ocean. A critical assessment of published distribution records and additional identifications based on cox1, psbA and rbcL genetic markers confirm the presence of D. acutiloba at a number of locations in the Indo-Pacific as well as three locations along the Israeli coastline in the south-eastern Mediterranean Sea. The close genetic affinity between introduced specimens and those from a population in Hurghada (Red Sea) strongly suggests an introduction via the Suez Canal. The occurrence of D. acutiloba in these regions is further supported by predictions made by correlative ecological niche models (ENMs), which show high suitability values in the northern Red Sea and the Levantine Basin. In contrast, environmental conditions in the western Mediterranean and parts of the north-eastern Mediterranean are currently less favourable, as evidenced by the lower predicted probability of occurrence. Under future scenarios, the suitability of these regions increases. The further spread of D. acutiloba in the eastern Mediterranean seems imminent, and the species may even extend its range to the western Mediterranean Sea, depending on the climate change scenario considered. While D. acutiloba can be relatively easily distinguished from the majority of Mediterranean Dictyota species, it remains difficult to differentiate this species from narrow growth forms of D. dichotoma. Therefore, we recommend the use of molecular markers such as cox1, psbA and rbcL, to unequivocally identify this species and monitor its further spread.

ABSTRACT Analysis of trait trade-offs, through which physiological traits requiring common resources are ‘traded’ to optimize competitive advantage, provides a route to simplify and more readily understand the complexities of ecology. The concept of trait trade-offs has found favour in plankton research, especially directed at phytoplankton, defined here as phototrophs incapable of phagotrophy. Mixoplankton, defined as protists that combine phototrophy and phagotrophy, are now recognized as being widespread and significant members of the protist plankton community; many photoflagellate ‘phytoplankton’ are actually mixoplankton, as are many ‘(microbial) zooplankton’. Mixoplankton might be expected to be dominant, being able to exploit different trophic strategies while simultaneously eliminating competitors. That mixoplankton are not dominant suggests that physiological trait trade-offs erode their apparent competitive edge. We present a systematic analysis of potential trait trade-offs in phototrophic protists focused on mixoplankton. We find no clear evidence to support trait trade-off arguments in plankton research, except perhaps for acquired phototrophy in mixoplanktonic ciliates versus zooplanktonic ciliates. Our findings suggest that the presence of various mixoplankton throughout the surface ocean waters is most likely explained by factors other than trait trade-offs. Diversities in mixoplankton form and function thus reflect that evolution of these organisms from very different lineages, provide them with advantages to function competitively in mature ecosystems with complex trophic interplay. Indeed, the complexity of those lineages is inconsistent with core trait trade-off definitions; there is no single ancestral mixoplankton nor a common environment supporting trait-trade-off-directed evolution.

ABSTRACT A unicellular cyanobacterium, strain VI4D9, was isolated from thermophilic microbial mats thriving in a hot spring of the Ahousaht territory of Vancouver Island, Canada, and characterized using optical and electron microscopy, genome sequencing and cultivation approaches. The cells were elongated rods (5.1 µm in length and 1.2 µm in width, on average). Their UV visible absorption spectra revealed that they contain chlorophyll a, phycocyanin and carotenoids. Transmission electron microscopy showed the presence of thylakoids concentrated on one side of the cells. The strain grew within a temperature range of 37–50°C, with an optimum growth at 45°C. Its genome had a size of 3 049 282 bp and a DNA G + C content of 51.8 mol%. The cells contained numerous intracellular spherical granules easily visible under scanning electron microscopy. Energy dispersive X-ray spectroscopy revealed that these granules were made of Ca-, Ba- and Sr-containing carbonates. A phylogenetic 16S rRNA gene tree robustly placed this strain as sister to several environmental sequences and the described species Gloeomargarita lithophora, also characterized by the possession of intracellular carbonate inclusions. We consider strain VI4D9 to represent a new Gloeomargarita species based on its marked phenotypic differences with G. lithophora, notably, its thermophilic nature and different thylakoid organization, therefore we propose the name Gloeomargarita ahousahtiae sp. nov. The type strain is VI4D9 (Culture Collection of Algae and Protozoa strain 1472/1; Laboratorio de Algas Continentales Mexico strain LAC 140). Gloeomargarita ahousahtiae is the second species described within the recently discovered order Gloeomargaritales.

ABSTRACT Among marine primary producers, macroalgae support complex and productive coastal food webs, but coastal primary production relies on terrigenous inputs and remineralized organic matter which both vary seasonally. An approach combining stable isotope and biochemical analyses enables a better characterization of macroalgae specificities and highlights environmental influences on their chemical signature. This study compared the isotopic signature and biochemical composition of 22 Mediterranean macroalgae belonging to Rhodophyta (red algae), Phaeophyceae (brown algae) and Chlorophyta (green algae) between March and November 2010 to capture the differences in species chemical signatures potentially driven by metabolic traits or environmental drivers. Carbon stable isotope values were evidenced as a good proxy of specific carbon metabolism: low values observed in red algae could be related to the reported absence of carbon concentrating mechanisms (CCMs) in this group while higher values were driven by strong CCM activity in green algae. Biochemical patterns also differed between groups: soluble carbohydrates were a major component for red algae, while lipids and proteins dominated in brown algae, and insoluble carbohydrate concentrations were high in green algae. Variation within species across two collection times could be related to environmental changes and algal metabolism. δ15N values confirm the efficiency of this parameter as a proxy of the impact of human influence in the Bay of Marseille.

ABSTRACT The characteristics of two new benthic species of Pyramimonas isolated from coral rubble have been determined. One, P. superba sp. nov., most similar to P. lunata, has a morphological character suite and molecular phylogenetic signal supporting it as a member of the subgenus Trichocystis, despite its uniqueness in producing mucilage. The other, P. lamellipunctata sp. nov., also shows phylogenetic affiliation with the subgenus Trichocystis using partial SSU data. However, this second species aligns with muciferous punctate species using rbcL data, although the clade is problematic as it includes the genus Pterosperma. Morphologically, cells of P. lamellipunctata share many features with muciferous punctate species but also exhibit important disparities with this group, most notably a lack of mucilage, suggesting that they form a unique lineage. HIGHLIGHTS ● Two new species of Pyramimonas are indicated as members of the subgenus Trichocystis using SSU data.● rbcL data only supports this association for one of the new species and affiliates the other with muciferous species of the subgenus Punctatae.● Morphological data only partially supports the second affiliation with punctate species, thereby inferring a unique lineage.

ABSTRACT Cyanobacterial blooms are widely known to cause problems in the aquatic environment, and their appearance has become more frequent due to global warming. Microcystis is one of the most widespread and dominant bloom-forming cyanobacterial genera, largely because Microcystis has the ability to control its buoyancy. A buoyancy experiment conducted on Microcystis sp. isolated from cyanobacterial blooms in Lake Senba, Japan, showed that buoyancy could be controlled using a combination of preculture under the dark conditions and the addition of powdered tightly bound extracellular polysaccharides (TB-EPS) and metal cations (Ca2+ and Mg2+). Preculture under the dark conditions, 96 h in length were the most effective treatment to reduce the cellular carbohydrate content of Microcystis and simultaneously increase its buoyancy. The addition of TB-EPS, Ca2+ and Mg2+ ions increased the colony size of Microcystis and enhanced buoyancy in precultures under both dark and light conditions. Thus, the buoyancy of Microcystis can be controlled by reducing its cellular carbohydrate content by preculturing it in dark conditions for 96 h and increasing the colony size with the addition of 100 mg l−1 EPS, 80 mg l−1 Ca2+ and 80 mg l−1 Mg2+. This study contributes to establishing a novel removal method for cyanobacterial blooms dominated by Microcystis, especially in water treatment facilities.