Enhancing biomass and lipid accumulation by a novel microalga for unsterilized piggery biogas slurry remediation.

The cost and efficiency of an algal-BS treatment system are determined by the specific microalgal species and BS pretreatment method. This study examines the growth of a novel algae Chlorella sp. YSD-2 and the removal of nutrients from the BS using different pretreatment methods, including dilution ratio and sterilization. The highest biomass production (1.84 g L-1) was achieved in the 1:2 unsterilized biogas slurry, which was 2.03 times higher than that in the sterilized group, as well as higher lipid productivity (17.29 mg L-1 d-1). Nevertheless, the sterilized biogas slurry at a 1:1 dilution ratio exhibited the most notable nutrient-removal efficiency, with COD at 71.97%, TP at 91.32%, and TN at 88.80%. Additionally, the analysis of 16S rRNA sequencing revealed a significant alteration in the indigenous bacterial composition of the biogas slurry by microalgal treatment, with Proteobacteria and Cyanobacteria emerging as the predominant phyla, and unidentified_Cyanobacteria as the primary genus. These findings suggest that Chlorella sp. YSD-2 exhibits favorable tolerance and nutrient-removal capabilities in unsterilized, high-strength biogas slurry, along with high productivity of biomass and lipids. Consequently, these results offer a theoretical foundation for the development of an efficient and economically viable treatment method for algal-BS.

Impact of high Spirulina diet, extruded or supplemented with enzymes, on blood cells, systemic metabolites, and hepatic lipid and mineral profiles of broiler chickens.

The impact of 15% dietary inclusion of Spirulina (Arthrospira platensis) in broiler chickens was explored, focusing on blood cellular components, systemic metabolites and hepatic lipid and mineral composition. From days 14 to 35 of age, 120 broiler chickens were divided and allocated into four dietary treatments: a standard corn and soybean meal-based diet (control), a 15% Spirulina diet, a 15% extruded Spirulina diet, and a 15% Spirulina diet super-dosed with an enzyme blend (0.20% porcine pancreatin plus 0.01% lysozyme). The haematological analysis revealed no significant deviations (p > 0.05) in blood cell counts across treatments, suggesting that high Spirulina inclusion maintains haematological balance. The systemic metabolic assessment indicated an enhanced antioxidant capacity in birds on Spirulina diets (p < 0.001), pointing toward a potential reduction in oxidative stress. However, the study noted a detrimental impact on growth performance metrics, such as final body weight and feed conversion ratio (both p < 0.001), in the Spirulina-fed treatments, with the super-dosed enzyme blend supplementation failing to alleviate these effects but with extrusion mitigating them. Regarding hepatic composition, birds on extruded Spirulina and enzyme-supplemented diets showed a notable increase in n-3 fatty acids (EPA, DPA, DHA) (p < 0.001), leading to an improved n-6/n-3 PUFA ratio (p < 0.001). Despite this positive shift, a reduction in total hepatic lipids (p = 0.003) was observed without a significant change in cholesterol levels. Our findings underscore the need for further exploration into the optimal inclusion levels, processing methods and potential enzymatic enhancements of Spirulina in broiler diets. Ultimately, this research aims to strike a balance between promoting health benefits and maintaining optimal growth performance in poultry nutrition.

Enhancing microalgae biomass production: Exploring improved scraping frequency in a hybrid cultivation system.

Recently, hybrid systems, such as those incorporating high-rate algal ponds (HRAPs) and biofilm reactors (BRs), have shown promise in treating domestic wastewater while cultivating microalgae. In this context, the objective of the present study was to determine an improved scraping frequency to maximize microalgae biomass productivity in a mix of industrial (fruit-based juice production) and domestic wastewater. The mix was set to balance the carbon/nitrogen ratio. The scraping strategy involved maintaining 1 cm wide stripes to retain an inoculum in the reactor. Three scraping frequencies (2, 4, and 6 days) were evaluated. The findings indicate that a scraping frequency of each 2 days provided the highest biomass productivity (18.75 g total volatile solids m-2 d-1). The species' behavior varied with frequency: Chlorella vulgaris was abundant at 6-day intervals, whereas Tetradesmus obliquus favored shorter intervals. Biomass from more frequent scraping demonstrated a higher lipid content (15.45%). Extrapolymeric substance production was also highest at the 2-day frequency. Concerning wastewater treatment, the system removed 93% of dissolved organic carbon and ∼100% of ammoniacal nitrogen. Combining industrial and domestic wastewater sources to balance the carbon/nitrogen ratio enhanced treatment efficiency and biomass yield. This study highlights the potential of adjusting scraping frequencies in hybrid systems for improved wastewater treatment and microalgae production.

Prefeeding of Clarias gariepinus with Spirulina platensis counteracts petroleum hydrocarbons-induced hepato- and nephrotoxicity.

Petroleum aromatic hydrocarbons are considered one of the most dangerous aquatic pollutants due to their widespread across water bodies, persistence, and extension to the food chain. To our knowledge, there hasn't been any research investigating the hepatorenoprotective effects of Spirulina platensis (SP) against toxicity induced by these environmental toxicants in fish. Thus, we decided to explore its potential safeguarding against benzene and toluene exposure in adult Clarias gariepinus. To achieve this objective, fish were divided into five groups (60 per group; 20 per replicate). The first group served as a control. The second and third groups were intoxicated with benzene and toluene at doses of 0.762 and 26.614 ng/L, respectively for 15 days. The fourth and fifth groups (SP + benzene and SP + toluene, respectively) were challenged with benzene and toluene as previously mentioned following dietary inclusion of SP at a dose of 5 g/kg diet for 30 days. The marked increase in liver metabolizing enzymes, glucose, total protein, albumin, globulin, albumin/globulin ratio, and creatinine confirmed the hepato- and nephrotoxic impacts of benzene and toluene. These outcomes were coupled with cytopathological affections and excessive collagen deposition. The incorporation of SP in ration formulation, on the contrary, restored the previously mentioned toxicological profile due to its antioxidant and cytoprotective attributes. Regardless of SP intervention, the renal tissues still displayed histo-architectural lesions, because of insufficient dose and timeframe. Additional research will be required to identify the ideal SP remediation regimen.

Features of the microalga Raphidocelis subcapitata: physiology and applications.

The microalga Raphidocelis subcapitata was isolated from the Nitelva River (Norway) and subsequently deposited in the collection of the Norwegian Institute of Water Research as "Selenastrum capricornutum Printz". This freshwater microalga, also known as Pseudokirchneriella subcapitata, acquired much of its notoriety due to its high sensitivity to different chemical species, which makes it recommended by different international organizations for the assessment of ecotoxicity. However, outside this scope, R. subcapitata continues to be little explored. This review aims to shed light on a microalga that, despite its popularity, continues to be an "illustrious" unknown in many ways. Therefore, R. subcapitata taxonomy, phylogeny, shape, size/biovolume, cell ultra-structure, and reproduction are reviewed. The nutritional and cultural conditions, chronological aging, and maintenance and preservation of the alga are summarized and critically discussed. Applications of R. subcapitata, such as its use in aquatic toxicology (ecotoxicity assessment and elucidation of adverse toxic outcome pathways) are presented. Furthermore, the latest advances in the use of this alga in biotechnology, namely in the bioremediation of effluents and the production of value-added biomolecules and biofuels, are highlighted. To end, a perspective regarding the future exploitation of R. subcapitata potentialities, in a modern concept of biorefinery, is outlined. KEY POINTS: • An overview of alga phylogeny and physiology is critically reviewed. • Advances in alga nutrition, cultural conditions, and chronological aging are presented. • Its use in aquatic toxicology and biotechnology is highlighted.

The cadmium tolerance and bioaccumulation mechanism of Tetratostichococcus sp. P1: insight from transcriptomics analysis.

Cadmium (Cd) has become a severe issue in relatively low concentration and attracts expert attention due to its toxicity, accumulation, and biomagnification in living organisms. Cd does not have a biological role and causes serious health issues. Therefore, Cd pollutants should be reduced and removed from the environment. Microalgae have great potential for Cd absorption for waste treatment since they are more environmentally friendly than existing treatment methods and have strong metal sorption selectivity. This study evaluated the tolerance and ability of the microalga Tetratostichococcus sp. P1 to remove Cd ions under acidic conditions and reveal mechanisms based on transcriptomics analysis. The results showed that Tetratostichococcus sp. P1 had a high Cd tolerance that survived under the presence of Cd up to 100 µM, and IC50, the half-maximal inhibitory concentration value, was 57.0 µM, calculated from the change in growth rate based on the chlorophyll content. Long-term Cd exposure affected the algal morphology and photosynthetic pigments of the alga. Tetratostichococcus sp. P1 removed Cd with a maximum uptake of 1.55 mg g-1 dry weight. Transcriptomic analysis revealed the upregulation of the expression of genes related to metal binding, such as metallothionein. Group A, Group B transporters and glutathione, were also found upregulated. While the downregulation of the genes were related to photosynthesis, mitochondria electron transport, ABC-2 transporter, polysaccharide metabolic process, and cell division. This research is the first study on heavy metal bioremediation using Tetratostichococcus sp. P1 and provides a new potential microalga strain for heavy metal removal in wastewater.[Figure: see text]Abbreviations:BP: Biological process; bZIP: Basic Leucine Zipper; CC: Cellular component; ccc1: Ca (II)-sensitive cross complementary 1; Cd: Cadmium; CDF: Cation diffusion facilitator; Chl: Chlorophyll; CTR: Cu TRansporter families; DAGs: Directed acyclic graphs; DEGs: Differentially expressed genes; DVR: Divinyl chlorophyllide, an 8-vinyl-reductase; FPN: FerroportinN; FTIR: Fourier transform infrared; FTR: Fe TRansporter; GO: Gene Ontology; IC50: Growth half maximal inhibitory concentration; ICP: Inductively coupled plasma; MF: molecular function; NRAMPs: Natural resistance-associated aacrophage proteins; OD: Optical density; RPKM: Reads Per Kilobase of Exon Per Million Reads Mapped; VIT1: Vacuolar iron transporter 1 families; ZIPs: Zrt-, Irt-like proteins.

Bicistronic expression of nuclear transgenes in Chlamydomonas reinhardtii.

In eukaryotic organisms, proteins are typically translated from monocistronic messenger RNAs containing a single coding sequence (CDS). However, recent long transcript sequencing identified 87 nuclear polycistronic mRNAs in Chlamydomonas reinhardtii natively carrying multiple co-expressed CDSs. In this study, we investigated the dynamics of 22 short intergenic sequences derived from these native polycistronic loci by their application in genetic constructs for synthetic transgene expression. A promising candidate sequence was identified based on the quantification of transformation efficiency and expression strength of a fluorescence reporter protein. Subsequently, the expression of independent proteins from one mRNA was verified by cDNA amplification and protein molecular mass characterization. We demonstrated engineered bicistronic expression in vivo to drive successful co-expression of several terpene synthases with the selection marker aphVIII. Bicistronic transgene design resulted in significantly increased (E)-α-bisabolene production of 7.95 mg L-1 from a single open reading frame, 18.1× fold higher than previous reports. Use of this strategy simplifies screening procedures for identification of high-level expressing transformants, does not require the application of additional fluorescence reporters, and reduces the nucleotide footprint compared to classical monocistronic expression cassettes. Although clear advantages for bicistronic transgene expression were observed, this strategy was found to be limited to the aphVIII marker, and further studies are necessary to gain insights into the underlying mechanism that uniquely permits this co-expression from the algal nuclear genome.

Complete mitochondrial genome of an oleaginous microalga Vischeria punctata (Eustigmatophyceae: Chlorobotryaceae) and phylogenetic analysis.

Vischeria punctata, as first described by Vischer in 1945, is a member of the family Chlorobotryaceae, within the order Eustigmatales. This species is recognized for its potential as a source of biofuels and other high-value products. In the present investigation, the whole genome of V. punctata was sequenced utilizing the Illumina HiSeq 4000 platform, enabling the assembly and annotation of its complete mitochondrial genome. The resulting circular genome spans 41,528 base pairs (bp) with a guanine-cytosine (GC) content of 27.3%. This genome encompasses 36 protein-coding genes, alongside 28 transfer RNA (tRNA), and three ribosomal RNA (rRNA) genes. The evolutionary trajectory of V. punctata was further explored by constructing a phylogenetic tree derived from the mitochondrial 33 gene dataset of 16 Ochrophyta species. Comparative analysis reveals that V. punctata bears closer ties to Vischeria sp. CAUP Q202 than to Vischeria stellata strain SAG 33.83, suggesting shared evolutionary pathways and phenotypic traits. This investigation constitutes the inaugural study into the mitochondrial evolution and phylogenetic patterning of the mitogenome in V. punctata. The outcomes from this research bolster our understanding of the genetic diversity and evolutionary processes within the class Eustigmatophyceae. In particular, the mitochondrial genome of V. punctata serves as a valuable resource in elucidating these aspects.

Chromosome-scale genome assembly of an acidophilic microalga Tetratostichococcus sp. P1 isolated from a tropical peatland in Malaysia.

Tetratostichococcus sp. P1 shows an acidophilic phenotype which could allow mass-scale monoculture of this green microalga without severe contamination by environmental microorganisms. In this study, we report a chromosome-scale genome assembly for Tetratostichococcus sp. P1.

Microalgae as an Alternative Mineral Source in Poultry Nutrition.

This review explores the potential of microalgae as a sustainable and nutritionally rich alternative for mineral supplementation in poultry diets, addressing both the opportunities and challenges in this emerging field. Poultry nutrition, pivotal to the health and productivity of birds, traditionally relies on inorganic and organic mineral sources which, while effective, raise environmental and economic concerns. Microalgae offer a promising solution with their high contents of essential minerals, proteins, vitamins, and bioactive compounds. This review delves into the nutritional profiles of various microalgae, highlighting their rich contents of minerals which are crucial for physiological processes in poultry. It examines the bioavailability of these minerals and their impact on poultry health and productivity. Furthermore, it evaluates the environmental sustainability of microalgae cultivation and acknowledges the challenges in using microalgae in poultry diets, particularly in terms of the economic viability of large-scale production and the consistency of nutrient composition. It discusses the importance of rigorous safety assessments and regulatory compliance, given the potential risks of toxins and heavy metals. Overall, this analysis aims to provide a clear understanding of the role microalgae could play in poultry nutrition and address sustainability challenges in animal agriculture while also considering future perspectives and advancements needed in this field.

Biodegradation of benzo[a]pyrene by a marine Chlorella vulgaris LH-1 with heterotrophic ability.

In this study, a microalga, Chlorella vulgaris LH-1, with heterotrophic ability to degrade BaP was explored. The effect of BaP concentration on microalga growth was investigated, and the possible biodegradation mechanism of BaP was proposed. Results showed that low BaP concentration (<5 mg/L) had less negative influence on the growth of this microalga under mixotrophic condition, but high BaP concentration (>5 mg/L) had a significant inhibitory effect on its growth. During heterotrophic cultivation, low BaP concentration (<20 mg/L) promoted the growth of C. vulgaris LH-1, whereas high BaP concentration (>20 mg/L) inhibited its growth significantly. The degradation rates of mixotrophic and heterotrophic C. vulgaris LH-1 were 62.56 %-74.13 % and 52.07 %-71.67 %, respectively, when the BaP concentration ranged from 0.5 mg/L to 2 mg/L. The expression of functional enzyme genes of C. vulgaris LH-1 such as phenol 2-monooxygenase activity, protocatechuate 3,4-dioxygenase activity, catechol 1,2-dioxygenase activity, styrene degradation, and benzoate degradation were upregulated in the process of BaP degradation. C. vulgaris LH-1 may degrade BaP by monooxygenase and dioxygenase simultaneously. The degradation of BaP by this microalga under mixotrophic condition goes through the degradation pathway of phthalic acid, whereas it goes through the degradation pathway of benzoic acid under heterotrophic condition.

The first arctic strain of Rhizochromulina: Morphology, ultrastructure, and position in the evolutionary tree of the order Rhizochromulinales (Heterokontophyta, Dictyochophyceae).

Rhizochromulina marina is a unicellular amoeboid alga capable of forming flagellate cells; it is a single validly named species in the genus. Besides, there are numerous environmental sequences and undescribed strains designated as Rhizochromulina sp. or R. marina. The biogeography of the genus is understudied: rhizochromulines from the Indian, Southern, and Arctic Oceans are unknown. Here, we present the description of Rhizochromulina sp. B44, which was for the first time isolated from an arctic habitat. Biofilms of this microalga grow at the bottom of a culture vessel, where neighbouring amoeboid cells form associations through a common network of pseudopodia, i.e. meroplasmodia. Pseudopodia branch, anastomose mainly during meroplasmodia formation, and are supported by microtubules that arise from the perinuclear zone. Actin filaments are localized in the cytoplasm and can be revealed only near the bases of pseudopodia. We succeeded in inducing the transformation of amoeboid cells into flagellates using a prolonged agitation of cultures. Morphological and molecular analyses revealed that the studied strain is most closely related to the type strain of R. marina. At the same time, 18S rDNA sequences of early branching-off rhizochromulinids differ significantly from Rhizochromulina sp. B44, suggesting a high divergence at the genus level.

Pioneering DNA assembling techniques and their applications in eukaryotic microalgae.

Assembling DNA fragments is a fundamental manipulation of cloning microalgal genes and carrying out microalgal synthetic biological studies. From the earliest DNA recombination to current trait and metabolic pathway engineering, we are always accompanied by homology-based DNA assembling. The improvement and modification of pioneering DNA assembling techniques and the combinational applications of the available assembling techniques have diversified and complicated the literature environment and aggravated our identification of the core and pioneering methodologies. Identifying the core assembling methodologies and using them appropriately and flourishing them even are important for researchers. A group of microalgae have been evolving as the models for both industrial applications and biological studies. DNA assembling requires researchers to know the methods available and their improvements and evolvements. In this review, we summarized the pioneering (core; leading) DNA assembling techniques developed previously, extended these techniques to their modifications, improvements and their combinations, and highlighted their applications in eukaryotic microalgae. We predicted that the gene(s) will be assembled into a functional cluster (e.g., those involving in a metabolic pathway, and stacked on normal microalgal chromosomes, their artificial episomes and looming artificial chromosomes. It should be particularly pointed out that the techniques mentioned in this review are classified according to the strategy used to assemble the final construct.

Microbial Diversity and Community Structure of Wastewater-Driven Microalgal Biofilms.

Dwindling water sources increase the need for efficient wastewater treatment. Solar-driven algal turf scrubber (ATS) system may remediate wastewater by supporting the development and growth of periphytic microbiomes that function and interact in a highly dynamic manner through symbiotic interactions. Using ITS and 16S rRNA gene amplicon sequencing, we profiled the microbial communities of four microbial biofilms from ATS systems operated with municipal wastewater (mWW), diluted cattle and pig manure (CattleM and PigM), and biogas plant effluent supernatant (BGE) in comparison to the initial inocula and the respective wastewater substrates. The wastewater-driven biofilms differed significantly in their biodiversity and structure, exhibiting an inocula-independent but substrate-dependent establishment of the microbial communities. The prokaryotic communities were comparable among themselves and with other microbiomes of aquatic environments and were dominated by metabolically flexible prokaryotes such as nitrifiers, polyphosphate-accumulating and algicide-producing microorganisms, and anoxygenic photoautotrophs. Striking differences occurred in eukaryotic communities: While the mWW biofilm was characterized by high biodiversity and many filamentous (benthic) microalgae, the agricultural wastewater-fed biofilms consisted of less diverse communities with few benthic taxa mainly inhabited by unicellular chlorophytes and saprophytes/parasites. This study advances our understanding of the microbiome structure and function within the ATS-based wastewater treatment process.

Ascorbate Peroxidase 2 (APX2) of Chlamydomonas Binds Copper and Modulates the Copper Insertion into Plastocyanin.

Recent phylogenetic studies have unveiled a novel class of ascorbate peroxidases called "ascorbate peroxidase-related" (APX-R). These enzymes, found in green photosynthetic eukaryotes, lack the amino acids necessary for ascorbate binding. This study focuses on the sole APX-R from Chlamydomonas reinhardtii referred to as ascorbate peroxidase 2 (APX2). We used immunoblotting to locate APX2 within the chloroplasts and in silico analysis to identify key structural motifs, such as the twin-arginine transport (TAT) motif for lumen translocation and the metal-binding MxxM motif. We also successfully expressed recombinant APX2 in Escherichia coli. Our in vitro results showed that the peroxidase activity of APX2 was detected with guaiacol but not with ascorbate as an electron donor. Furthermore, APX2 can bind both copper and heme, as evidenced by spectroscopic, and fluorescence experiments. These findings suggest a potential interaction between APX2 and plastocyanin, the primary copper-containing enzyme within the thylakoid lumen of the chloroplasts. Predictions from structural models and evidence from 1H-NMR experiments suggest a potential interaction between APX2 and plastocyanin, emphasizing the influence of APX2 on the copper-binding abilities of plastocyanin. In summary, our results propose a significant role for APX2 as a regulator in copper transfer to plastocyanin. This study sheds light on the unique properties of APX-R enzymes and their potential contributions to the complex processes of photosynthesis in green algae.

The Antiviral Potential of Algal Lectins.

Algae have emerged as fascinating subjects of study due to their vast potential as sources of valuable metabolites with diverse biotechnological applications, including their use as fertilizers, feed, food, and even pharmaceutical precursors. Among the numerous compounds found in algae, lectins have garnered special attention for their unique structures and carbohydrate specificities, distinguishing them from lectins derived from other sources. Here, a comprehensive overview of the latest scientific and technological advancements in the realm of algal lectins with a particular focus on their antiviral properties is provided. These lectins have displayed remarkable effectiveness against a wide range of viruses, thereby holding great promise for various antiviral applications. It is worth noting that several alga species have already been successfully commercialized for their antiviral potential. However, the discovery of a diverse array of lectins with potent antiviral capabilities suggests that the field holds immense untapped potential for further expansion. In conclusion, algae stand as a valuable and versatile resource, and their lectins offer an exciting avenue for developing novel antiviral agents, which may lead to the development of cutting-edge antiviral therapies.

[Preparation, characterization and activity evaluation of Spirulina-chitooligosaccharides capable of inhibiting biofilms].

The biofilms formed by pathogenic microorganisms seriously threaten human health and significantly enhance drug resistance, which urgently call for developing drugs specifically targeting on biofilms. Chitooligosaccharides extracted from shrimp and crab shells are natural alkaline oligosaccharides with excellent antibacterial effects. Nevertheless, their inhibition efficacy on biofilms still needs to be improved. Spirulina (SP) is a microalga with negatively charged surface, and its spiral structure facilitates colonization in the depth of the biofilm. Therefore, the complex of Spirulina and chitooligosaccharides may play a synergistic role in killing pathogens in the depth of biofilm. This research first screened chitooligosaccharides with significant bactericidal effects. Subsequently, Spirulina@Chitooligosaccharides (SP@COS complex was prepared by combining chitooligosaccharides with Spirulina through electrostatic adsorption. The binding of the complex was characterized by zeta potential, z-average size, and fluorescence labeling. Ultraviolet-visible spectroscopy (UV-Vis) showed the encapsulation efficiency and the drug loading efficiency reached up to 90% and 16%, respectively. The prepared SP@COS2 exhibited a profound synergistic inhibition effect on bacterial and fungal biofilms, which was mainly achieved by destroying the cell structure of the biofilm. These results demonstrate the potential of Spirulina-chitooligosaccharides complex as a biofilm inhibitor and provide a new idea for addressing the harm of pathogenic microorganisms.

The effect of Dunaliella tertiolecta supplementation on diet-induced obesity in UCP1-deficient mice.

We previously demonstrated that dietary supplementation with Dunaliella tertiolecta (DT) increases uncoupling protein 1 (UCP1) expression in brown adipose tissue (BAT) and improves diet-induced obesity (DIO) in C57BL/6 J mice at thermoneutrality (30 °C). Here, we investigated whether DT improves DIO in a thermoneutral UCP1-deficient (KO) animal. KO mice were fed a high-fat diet supplemented with DT for 12 weeks. Compared to control group without DT, body weight was significantly reduced in DT group with no difference in food intake. Dunaliella tertiolecta-supplemented mice exhibited lower adiposity and well-maintained multilocular morphology in BAT, in which a significant increase in gene expression of PR domain containing 16 was detected in DT group compared to control group. Moreover, increase in UCP2 level and/or decrease in ribosomal protein S6 phosphorylation were detected in adipose tissues of DT group relative to control group. These results suggest that DT supplementation improves DIO by stimulating UCP1-independent energy dissipation at thermoneutrality.