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Comparison of P. soloecismus growth in different salinity concentrations, upon shifting from a starting concentration of 35 ppt (100% Instant Ocean seawater mix in f/2, ppt = parts per thousand) to each salinity shown in the legend. A) Growth observed as an increase in optical density (750 nm), B) Growth rates at the exponential phase for each salinity except 140 ppt, which grew too poorly to calculate a growth rate. Cells were grown in Erlenmeyer flasks in a growth chamber at 1% CO 2 and 25 °C
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Photosynthetic microbes are considered promising biofactories for transforming inorganic carbon from the atmosphere into a renewable source of chemicals and precursors of industrial interest; however, there continues to be a need for strains that demonstrate high productivity, environmental robustness, and the potential to be genetically manipulate...
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... replenishing after biomass harvesting [58,59]. Therefore, we assessed the native tolerance of P. soloecismus to changes in salinity, testing a range from 0.35 to 140 ppt. Apart from any adaptation that may have occurred during the 13 day experiment, these results describe the tolerance to "shocking" the culture with a variety of salinities. Fig. 4A shows the growth of P. soloecismus in seven different salinities, and Fig. 4B shows growth rates calculated during exponential growth ( Supplementary Fig. 4) after the respective lag for each condition, except 140 ppt (rate values and statistics in Supplementary Table 3). Although OD 750 appeared to increase exponentially for days 3-5 ...
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... native tolerance of P. soloecismus to changes in salinity, testing a range from 0.35 to 140 ppt. Apart from any adaptation that may have occurred during the 13 day experiment, these results describe the tolerance to "shocking" the culture with a variety of salinities. Fig. 4A shows the growth of P. soloecismus in seven different salinities, and Fig. 4B shows growth rates calculated during exponential growth ( Supplementary Fig. 4) after the respective lag for each condition, except 140 ppt (rate values and statistics in Supplementary Table 3). Although OD 750 appeared to increase exponentially for days 3-5 for the 0.35 ppt and 140 ppt cultures, the overall optical densities for these ...
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... from any adaptation that may have occurred during the 13 day experiment, these results describe the tolerance to "shocking" the culture with a variety of salinities. Fig. 4A shows the growth of P. soloecismus in seven different salinities, and Fig. 4B shows growth rates calculated during exponential growth ( Supplementary Fig. 4) after the respective lag for each condition, except 140 ppt (rate values and statistics in Supplementary Table 3). Although OD 750 appeared to increase exponentially for days 3-5 for the 0.35 ppt and 140 ppt cultures, the overall optical densities for these cultures were so low that we did not calculate growth rates for these days. ...
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... while a relatively tight range of performance was observed between 3.5 and 70 ppt, a large drop in growth was observed both from 3.5 ppt to 0.35 ppt and from 70 ppt to 140 ppt (Fig. 4A). Notably, after 10 days of incubation, the 0.35 ppt culture began to grow, although the growth rate was significantly lower than the 35 ppt growth rate (p < 0.001, Supplementary Table 3). Meanwhile, the 140 ppt culture did not increase in OD during the time frame of the experiment. Overall, these results show that P. soloecismus is ...
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... The genus Picochlorum is characterized by the following five morphological and eco-physiological characteristics (Henley et al., 2004;Temraleeva et al., 2022): (1) they are typically small, ranging from 1 to 4 µm in diameter; (2) They have one nucleus, one mitochondrion, and one lateral chloroplast (containing chlorophylls a and b, but without pyrenoids); (3) Generally, they do not have flagella; a newly described strain exceptionally showed flagella (Pang et al., 2022); (4) The cells reproduce by autosporulation into two or more daughter cells; (5) The species inhabits marine environments. In addition, Picochlorum species are known for their high photosynthetic efficiency, fast growth rate, and ability to produce large amounts of lipids (Dahlin et al., 2019;Gonzalez-Esquer et al., 2019;Krishnan et al., 2021;Zhu & Dunford, 2013). Because of these characteristics, they have potential applications in biotechnology and bioenergy and have been the focus of extensive research, especially in biodiesel production (Chakravarty & Mallick, 2022;Dogaris et al., 2019;Foflonker et al., 2018;Gonzalez-Esquer et al., 2018;Goswami et al., 2022). ...
... The strain showed optimal growth at a temperature of 40°C while being able to withstand peak temperatures up to 47.5°C under fluctuating diel temperature regimes (Barten et al., 2021). Picochlorum species have received increasing interest in research in recent years, due to their relatively high growth rates, their ability to survive under dynamic cultivation conditions, and their ability to tolerate high light intensities, temperatures, and salinity levels (Weissman et al., 2018;Gonzalez-Esquer et al., 2019;Barten et al., 2021;Krishnan et al., 2021). As a result of its thermo-tolerance, the cultivation of Picochlorum sp. may enable a reduction in production costs of more than 25% (Barten et al., 2021). ...
Microalgae are a promising renewable feedstock that can be produced on non-arable land using seawater. Their biomass contains proteins, lipids, carbohydrates, and pigments, and can be used for various biobased products, such as food, feed, biochemicals, and biofuels. For such applications, the production costs need to be reduced, for example, by improving biomass productivity in photobioreactors. In this study, Picochlorum sp. (BPE23) was cultivated in a prototype of a novel outdoor V-shaped photobioreactor on Bonaire (12°N, 68°W). The novel photobioreactor design was previously proposed for the capture and dilution of sunlight at low-latitude locations. During several months, the biomass productivity of the local thermotolerant microalgae was determined at different dilution rates in continuous dilution and batch dilution experiments, without any form of temperature control. Reactor temperatures increased to 35°C–45°C at midday. In the continuous dilution experiments, high average biomass productivities of 28–31 g m⁻² d⁻¹ and photosynthetic efficiencies of 3.5%–4.3% were achieved. In the batch dilution experiments, biomass productivities were lower (17–23 g m⁻² d⁻¹), as microalgal cells likely experienced sudden light and temperature stress after daily reactor dilution. Nonetheless, dense cultures were characterized by high maximum photosynthetic rates, illustrating the potential of Picochlorum sp. for fast growth under outdoor conditions.
... The chloroplast and nucleus were colored blue and red, respectively (Figure 1j,k). In previous studies, a TEM image analysis revealed the organelle morphology of lipid bodies, starch granules, thylakoids, and mitochondria in the genus Picochlorum [21,25,29]. Lipid bodies, which were stained with Nile Red and are indicated by white arrows, exhibited a light green hue (Figure 1l,m). ...
Microalgae are highly valued for their rapid biomass production and metabolite synthesis, as well as their abundance of beneficial compounds. They have a variety of applications, including serving as the primary ingredient in biofuels, functional foods, and cosmetics. The genus Picochlorum, which was established to represent the unique characteristics of “Nannochloris-like” algae, exhibits rapid growth and a high salt tolerance. The morphology, molecular phylogeny, and fatty acid composition of an unspecified Picochlorum strain KCTC AG61293 found in Korean coastal waters were investigated. The strain exhibited a unique cell morphology and reproduction type compared to other Picochlorum species, as determined using light microscopy, fluorescence microscopy, and field emission scanning electron microscopy (FE-SEM). The vegetative cells were elongated and cylindrical in shape, underwent binary fission, and possessed a parietal chloroplast. A molecular phylogenetic analysis using nuclear small subunit ribosomal RNA sequences showed that Picochlorum sp. (KCTC AG61293) belongs to the Picochlorum clade and is closely related to the genus Nannochloris. Compared to other reference species, Picochlorum sp. (KCTC AG61293) had higher levels of saturated fatty acids (SFAs) and alpha-linolenic acid (ALA). The increased levels of SFAs and ALA suggest that Picochlorum sp. (KCTC AG61293) may be a promising candidate for biofuel production and other industrial uses.
... In microalgae F, the cells were unicellular and spherical with a nucleus, 8 to 10 μm diameter in size without flagella and pyrethroids, which were characteristics of Picochlorum spp. Microalgae F had quite distinct features in terms of size from other Picochlorum species, such as P. soloecismus and P. maculatum [24]. In the microalgae G, the cells were highly green chloroplasts, a pyrenoid inside the chloroplast, and a sheath wall, characteristic of Tetraselmis spp. ...
Algal bioprospecting in ecosystems leads to exploring native microalgae and the competency evaluation of economically producing lipids as biofuel or nutritional applications. In this study, ten microalgae species were screened from the saline water lake. Chlorella vulgaris, Chlorella sorokiniana, Chlamydomonas raudensis, Chlamydomonas hedleyi, Dunaliella salina, Picochlorum bazangan sp. nov., Tetraselmis bazangan sp. nov., Haematococcus lacustris, Nannochloropsis oceanic, and Scenedesmus rubescens were isolated and identified using 18SrDNA and tufA markers. Biodiesel potentials were assayed by the determination of biomass productivity, biochemical components, fatty acid profile, and biodiesel properties. The results showed that the maximum biomass yield (1.22 g/L) belonged to C. vulgaris. The highest protein, carbohydrate, chlorophyll, and carotenoid content were recorded in C. vulgaris, C. raudensis, C. sorokiniana, and D. salina, respectively. N. oceanica accumulated high lipid content and omega-3 fractions (31.09%). However, C. hedleyi had the highest lipid productivity (11.64 g/L/day) compared to other microalgae. The best species for biodiesel production was C. vulgaris, with a specific growth rate of 0.36 day⁻¹, lipid productivity of 7.45 g/L/day, and C16-C18 fatty acid profile of 78.3%. The microalgae C. vulgaris had appropriate biodiesel properties of low viscosity (4.49), high cetane number (55.38), and relatively low cloud point (4.98). Another choice was N. oceanic, with high lipid productivity, cetane number (59.79), oxidative stability (56.43), and low iodine value (47.11). Microalgae T. bazangan sp. nov. had a cetane number (55.24), low cloud point (4.71), and C16-C18 fatty acid profile of 82.34%. Accordingly, C. vulgaris, T. bazangan sp. nov., and N. oceanic can be considered potential species for biodiesel.
... Here, we describe depigmented strains of a remarkably fastgrowing, high-light tolerant microalga, Picochlorum celeri. The Picochlorum genus (Henley et al., 2004) has become the focus of several recent studies because of their high stress tolerances, as well as their small, compact genomes (Barten et al., 2020;Dahlin et al., 2019;Foflonker et al., 2016;Gonzalez-Esquer et al., 2019;Manjre et al., 2022;Weissman et al., 2018). P. celeri demonstrates broad halotolerance, an exceptional ability to thrive at high light intensities (>2000 μmol photons m À2 s À1 ), temperatures (>35 C), and salinities (2-3Â seawater), and can attain rapid doubling times of $2 h under optimal conditions . ...
High cellular pigment levels in dense microalgal cultures contribute to excess light absorption. To improve photosynthetic yields in the marine microalga Picochlorum celeri , CAS9 gene editing was used to target the molecular chaperone cpSRP43. Depigmented strains (>50% lower chlorophyll) were generated, with proteomics showing attenuated levels of most light harvesting complex (LHC) proteins. Gene editing generated two types of cpSRP43 transformants with distinct lower pigment phenotypes: (i) a transformant (Δ srp43 ) with both cp SRP43 diploid alleles modified to encode non‐functional polypeptides and (ii) a transformant (STR30309) with a 3 nt in‐frame insertion in one allele at the CAS9 cut site (non‐functional second allele), leading to expression of a modified cpSRP43. STR30309 has more chlorophyll than Δ srp43 but substantially less than wild type. To further decrease light absorption by photosystem I in STR30309, CAS9 editing was used to stack in disruptions of both LHCA6 and LHCA7 to generate STR30843, which has higher (5–24%) productivities relative to wild type in solar‐simulating bioreactors. Maximal productivities required frequent partial harvests throughout the day. For STR30843, exemplary diel bioreactor yields of ~50 g m ⁻² day ⁻¹ were attained. Our results demonstrate diel productivity gains in P. celeri by lowering pigment levels.
... For Class IV, Tetraselmis had the most relatively abundant ASVs, a genus that is known to have many euryhaline species (Fon-Sing and Borowitzka, 2016), but has not been well studied in the tropics (Arora et al., 2013). The most relatively abundant Trebouxiophyceae ASVs were related to Picochlorum soloecismus and Chlorella desiccata, also known to be halotolerant, but not previously reported from tropical environments (Gonzalez-Esquer et al., 2019;Borovsky et al., 2020). ...
Tropical environments with unique abiotic and biotic factors—such as salt ponds, mangroves, and coral reefs—are often in close proximity. The heterogeneity of these environments is reflected in community shifts over short distances, resulting in high biodiversity. While phytoplankton assemblages physically associated with corals, particularly their symbionts, are well studied, less is known about phytoplankton diversity across tropical aquatic environments. We assess shifts in phytoplankton community composition along inshore to offshore gradients by sequencing and analyzing 16S rRNA gene amplicons using primers targeting the V1-V2 region that capture plastids from eukaryotic phytoplankton and cyanobacteria, as well as heterotrophic bacteria. Microbial alpha diversity computed from 16S V1-V2 amplicon sequence variant (ASV) data from 282 samples collected in and around Curaçao, in the Southern Caribbean Sea, varied more within the dynamic salt ponds, salterns, and mangroves, compared to the seemingly stable above-reef, off-reef, and open sea environments. Among eukaryotic phytoplankton, stramenopiles often exhibited the highest relative abundances in mangrove, above-reef, off-reef, and open sea environments, where cyanobacteria also showed high relative abundances. Within stramenopiles, diatom amplicons dominated in salt ponds and mangroves, while dictyochophytes and pelagophytes prevailed above reefs and offshore. Green algae and cryptophytes were also present, and the former exhibited transitions following the gradient from inland to offshore. Chlorophytes and prasinophyte Class IV dominated in salt ponds, while prasinophyte Class II, including Micromonas commoda and Ostreococcus Clade OII, had the highest relative abundances of green algae in mangroves, above-reef, off-reef, and the open sea. To improve Class II prasinophyte classification, we sequenced 18S rRNA gene amplicons from the V4 region in 41 samples which were used to interrelate plastid-based results with information on uncultured prasinophyte species from prior 18S rRNA gene-based studies. This highlighted the presence of newly described Ostreococcus bengalensis and two Micromonas candidate species. Network analyses identified co-occurrence patterns between individual phytoplankton groups, including cyanobacteria, and heterotrophic bacteria. Our study reveals multiple uncultured and novel lineages within green algae and dictyochophytes in tropical marine habitats. Collectively, the algal diversity patterns and potential co-occurrence relationships observed in connection to physicochemical and spatial influences help provide a baseline against which future change can be assessed.
... This species was isolated from a culture of Nannochloropsis salina CCMP1776 after it outcompeted the native Nannochloropsis under heat stress [39]. P. soloecismus is relatively small, approximately 2.2 μm, though its size almost doubles under nitrogen stress [40]. Its 15 MB nuclear genome contains 12.1% 5mC CpG methylation with 48,667 CpG sites containing greater than 80% methylation [41]. ...
... (Henley et al. 2004). It has been reported that Picochlorum soloecismus could be a new platform for the production of renewable fuels (Gonzalez-Esquer et al. 2019). In addition, Picochlorum strain SENEW3 accumulated proline under high salt stress conditions through upregulation of one gene involved in proline synthesis (Foflonker et al. 2016). ...
Microalgae are considered the most promising source of renewable fuels, high-value bio-products and nutraceuticals. Potentially, microalgae can satisfy many global demands, but in large-scale cultivation the average productivity of most industrial strains is lower than maximal theoretical estimations, mainly due to sub-optimal growth conditions. Although microalgae have developed complex strategies to cope with environmental stresses, cultivation in outdoor photobioreactors is limited to few species and it is not yet sufficiently remunerative. Indeed, most microalgal species are very sensitive to environmental conditions, and changes in solar irradiation, temperature, and medium composition can drastically decrease biomass yield. Developing new strategies for improving algal tolerance to stress conditions is thus greatly desirable. One of the first responses that occur in both higher plants and microorganisms following the exposure to abiotic stress conditions, is an increased synthesis and accumulation of the amino acid proline. While the role of proline accumulation in stress adaptation is well-recognized in higher plants, in microalgae the implication of proline in stress tolerance still awaits full elucidation. In this review we summarize available data on proline metabolism under environmental stress in eukaryotic microalgae. Possible implications toward optimization of algal growth for biotechnological purposes are also discussed.
... % of the total phytoplankton, as revealed by high-throughput sequencing in the waters of a typical indoor industrial aquaculture system for farmed shrimp Litopenaeus vannamei. Species of the genus Picochlorum are small μm (Gonzalez-Esquer et al., 2019). Studies have shown that Picochlorum has high growth rates and biomass production and is rich in carotenoids, proteins and lipids (Zhu and Dunford, 2013;Unkefer et al., 2017;Dinesh Kumar et al., 2017;Gonzalez-Esquer et al., 2019). ...
... Species of the genus Picochlorum are small μm (Gonzalez-Esquer et al., 2019). Studies have shown that Picochlorum has high growth rates and biomass production and is rich in carotenoids, proteins and lipids (Zhu and Dunford, 2013;Unkefer et al., 2017;Dinesh Kumar et al., 2017;Gonzalez-Esquer et al., 2019). Picoplankton plays a vital role in marine food-web structure via energy transfer within the microbial loop. ...
Filter-feeder bivalves and phytoplankton are interdependent. Their interaction plays important role in estuarine and coastal ecosystem. The correlation between bivalve feeding and phytoplankton is highly species specificity and environment dependent. In the background of miniature and nondiatom trend of phytoplankton in coastal seawaters, how bivalve respond and how the response play roles in the phytoplankton community are poorly known. In the present study, by applying DNA metabarcoding approach based on plastid 23S rDNA, this question was addressed by comparing the phytoplankton composition in the seston and the stomach content of blood clam Tegillarca granosa sampled during the growth period from March to November 2020 in an experimental farm on tidal flat in Xiangshan Bay, East China Sea. The result showed that, a total of seven phyla, 55 genera and 73 species of phytoplankton were identified for all samples. Chlorophyta, Bacillariophyta, and Cyanobacteria were found to be three dominant phyla both in the stomach contents and seston. High diversity of pico-sized phytoplankton, which was easy overlooked by microscopy, was revealed both in seston and stomach contents. This result indicated that the clam was able to feed on the pico-sized algae. At the genus level, the most abundant genera were the pico-sized green alga Ostreococcus (6.12 %-67.88 %) in seston and Picochlorum (4.07 %-35.33 %) in the stomach contents. In addition, microalgae of high nutritional value showed trend of higher proportion in stomach contents than that in seston, especially in July and September when significant growth of T. granosa was observed during this period (the body size increased 155 %). Biodiversity of phytoplankton in the seston was totally higher than that in stomach content, however, the changes among the months showed respective trend. Especially in July when the biodiversity was the lowest in seston, that in the stomach content showed the highest. The results indicated that blood clam farming might influence the phytoplankton composition, including those of pico-sized level, although the particular species in seston were mainly correlating with the dominant environmental factors such as temperature, salinity, pH respectively. These results extend the understanding of roles that bivalve aquaculture may play in the changing of coastal phytoplankton community.
... These include species from the genera, Chlorella, Picochlorum, Scenedesmus, Cyclotela, Tetraselmus and Nanochloropsis ( Table 1). Many of these species have demonstrated long-term biomass production (>100 days) yields approaching 40 gdw/m 2 / day 22, (Huesemann et al., 2018;Dahlin et al., 2019;Gonzalez-Esquer et al., 2019;Aketo et al., 2020;Mucko et al., 2020;Cano et al., 2021;Krishnan et al., 2021). Averaged over a yearlong growing season algae having a growth rate of 40 gdw/m 2 /day could yield as much as 145 metric tons dw/ha/yr having an average energy density of 21 kJ/gdw. ...
... Recently, one genus of algae, Picochlorum, has emerged as one of the highest and most stable biomass producers (Dahlin et al., 2019;Gonzalez-Esquer et al., 2019;Mucko et al., 2020;Cano et al., 2021;Krishnan et al., 2021). Picochlorum are marine algae having a wide global distribution that tolerate high salinity and temperatures (40 C) well allowing for cultivation in a broad range of environments (Zhu et al., 2008;Dahlin et al., 2019;Krishnan et al., 2021). ...
... In contrast to efforts to improve biomass yields through alterations in light use efficiency, there have been limited efforts to improve algal biomass yields through manipulation of CBBC enzymes or by increasing carbon sink strength. There have, however, been substantial efforts to increase lipid accumulation in microalgae through genetic engineering and genome editing approaches often at a loss in biomass accumulation (Gonzalez-Esquer et al., 2019;Fayyaz et al., 2020;Ng et al., 2020;Shokravi et al., 2021). One approach to increase biomass accumulation through manipulation of the CBBC activity was achieved through enhancement of fructose 1,6-bisphosphate aldolase activity, an enzyme playing a major role in carbon flux control in the CBBC . ...
There is growing evidence that eukaryotic microalgae can become a more sustainable and profitable alternative than terrestrial crops to produce feed, fuels, and valuable coproducts. The major factor driving progress in algal biomass production is the potential of microalgae to produce substantially greater biomass per unit land area than terrestrial crops. To be financially feasible, however, current algal biomass yields must be increased. Given the fact that algal biomass production is in its infancy there exist multiple opportunities to improve biomass yields. For example, recent bioprospecting efforts have led to the identification of new microalgal strains having biomass yields that compete economically with plant biomass. Substantial increases in biomass yields have also been achieved using advanced genetic engineering approaches. Targeted improvements in photosynthetic efficiency have led to three-fold increases in algal biomass yields. One genetic tool that has seen limited application for algal biomass enhancement is advanced breeding genetics. The greater availability of algal genomes and recent advancements in breeding algae will further accelerate yield improvements. Genetic engineering strategies to increase biomass production will also be assisted by transcriptomic and metabolomic studies that help identify metabolic constraints that limit biomass production. In this review we assess some of the recent advances in algal strain selection, directed evolution, genetic engineering and molecular-assisted breeding that offer the potential for increased algal biomass production.
