The horizontal gene transfer of perchlorate reduction genomic island in three bacteria from an ecological niche.

Three new strains of dissimilatory perchlorate-reducing bacteria (DPRB), QD19-16, QD1-5, and P3-1, were isolated from an active sludge. Phylogenetic trees based on 16S rRNA genes indicated that QD19-16, QD1-5, and P3-1 belonged to Brucella, Acidovorax, and Citrobacter, respectively, expanding the distribution of DPRB in the Proteobacteria. The three strains were gram-negative and facultative anaerobes with rod-shaped cells without flagella, which were 1.0-1.6 µm long and 0.5-0.6 µm wide. The three DPRB strains utilized similar broad spectrum of electron donors and acceptors and demonstrated a similar capability to reduce perchlorate within 6 days. The enzyme activity of perchlorate reductase in QD19-16 toward chlorate was higher than that toward perchlorate. The high sequence similarity of the perchlorate reductase operon and chlorite dismutase genes in the perchlorate reduction genomic islands (PRI) of the three strains implied that they were monophyletic origin from a common ancestral PRI. Two transposase genes (tnp1 and tnp2) were found in the PRIs of strain QD19-16 and QD1-5, but were absent in the strain P3-1 PRI. The presence of fragments of IR sequences in the P3-1 PRI suggested that P3-1 PRI had previously contained these two tnp genes. Therefore, it is plausible to suggest that a common ancestral PRI transferred across the strains Brucella sp. QD19-16, Acidovorax sp. QD1-5, and Citrobacter sp. P3-1 through horizontal gene transfer, facilitated by transposases. These results provided a direct evidence of horizontal gene transfer of PRI that could jump across phylogenetically unrelated bacteria through transposase. KEY POINTS: • Three new DPRB strains can effectively remove high concentration of perchlorate. • The PRIs of three DPRB strains are acquired from a single ancestral PRI. • PRIs are incorporated into different bacteria genome through HGT by transposase.

Overexpression of plastid lipid-associated protein in marine diatom enhances the xanthophyll synthesis and storage.

Plastoglobules, which are lipoprotein structures surrounded by a single hydrophobic phospholipid membrane, are subcellular organelles in plant chromoplasts and chloroplasts. They contain neutral lipids, tocopherols, quinones, chlorophyll metabolites, carotenoids and their derivatives. Proteomic studies indicated that plastoglobules are involved in carotenoid metabolism and storage. In this study, one of the plastid lipid-associated proteins (PAP), the major protein in plastoglobules, was selected and overexpressed in Phaeodactylum tricornutum. The diameter of the plastoglobules in mutants was decreased by a mean of 19.2% versus the wild-type, while the fucoxanthin level was increased by a mean of 51.2%. All mutants exhibited morphological differences from the wild-type, including a prominent increase in the transverse diameter. Moreover, the unsaturated fatty acid levels were increased in different mutants, including an 18.9-59.3% increase in eicosapentaenoic acid content. Transcriptomic analysis revealed that PAP expression and the morphological changes altered xanthophyll synthesis and storage, which affected the assembly of the fucoxanthin chlorophyll a/c-binding protein and expression of antenna proteins as well as reduced the non-photochemical quenching activity of diatom cells. Therefore, metabolic regulation at the suborganelle level can be achieved by modulating PAP expression. These findings provide a subcellular structural site and target for synthetic biology to modify pigment and lipid metabolism in microalgae chassis cells.

Expression of the Vitreoscilla hemoglobin gene in Nannochloropsis oceanica regulates intracellular oxygen balance under high-light.

Nannochloropsis oceanica is widely used as a model photosynthetic chassis to produce fatty acids and carotenoid pigments. However, intense light typically causes excessive generation of reactive oxygen species (ROS) and photorespiration in microalgal cells, which results in decreased cell growth rate and unsaturated fatty acid content. In this study, the Vitreoscilla hemoglobin gene (vgb) was introduced into N. oceanica cells and expressed by using the light-harvesting complex promoter and its signal peptide. Compared with wild type (WT), the growth rate of transformants increased by 7.4%-18.5%, and the eicosapentaenoic acid content in an optimal transformant increased by 21.0%. Correspondingly, the intracellular ROS levels decreased by 56.9%-70.0%, and the catalase content in transformants was about 1.8 times that of WT. The photorespiration level of transformants was reduced by the measurement and calculation of the dissolved oxygen concentration under the condition of light-dark transition. The expression level of the key genes related to the photorespiration pathway in transformants was more than 80% lower than that in WT. These results indicated that Vitreoscilla hemoglobin could improve microalgal growth by reducing ROS damage and modulating photorespiration under stress conditions.

Naphthylacetic Acid and Tea Polyphenol Application Promote Biomass and Lipid Production of Nervonic Acid-Producing Microalgae.

Mychonastes afer HSO-3-1 is a potential producer of nervonic acid, which could be accumulated to 2-3% of dry cell weight. Improving the productivity of nervonic acid is critical to promote the commercialization of this product. In this study, 1-naphthylacetic acid (NAA) and tea polyphenol (TP) were selected as bioactive additives to stimulate the growth of M. afer. Supplementing NAA in the early growth stage and TP in the middle and late growth stage led to improved lipid accumulation in M. afer. The cultures supplemented with TP at the late growth stage maintained higher photosynthetic efficiency than the control groups without TP. Furthermore, the intracellular reactive oxygen species (ROS) accumulations in M. afer supplemented with 500 mg/L of TP was 63% lower than the control group. A linear relationship (R2= 0.899) between the values of Fv/Fm and ROS accumulation was established. We hypothesize supplement of bioactive additives at different growth stage could promote the cell growth rate and nervonic acid productivity of M. afer by retrieving intracellular ROS level. Further analysis of photosynthetic system II (PSII) protein in M. afer cultured in presence of NAA and TP indicated the levels of D1 and D2 proteins, the core skeleton proteins of PSII, showed 33.3 and 25.6% higher than the control group. CP43 protein, a critical module in PSII repair cycle, decreased significantly. These implied that TP possesses the function of slowing down the damage of PSII by scavenging excess intracellular ROS.

Biosynthesis of nervonic acid and perspectives for its production by microalgae and other microorganisms.

Nervonic acid (NA) is a major very long-chain monounsaturated fatty acid found in the white matter of mammalian brains, which plays a critical role in the treatment of psychotic disorders and neurological development. In the nature, NA has been synthesized by a handful plants, fungi, and microalgae. Although the metabolism of fatty acid has been studied for decades, the biosynthesis of NA has yet to be illustrated. Generally, the biosynthesis of NA is considered starting from oleic acid through fatty acid elongation, in which malonyl-CoA and long-chain acyl-CoA are firstly condensed by a rate-limiting enzyme 3-ketoacyl-CoA synthase (KCS). Heterologous expression of kcs gene from high NA producing species in plants and yeast has led to synthesis of NA. Nevertheless, it has also been reported that desaturases in a few plants can catalyze very long-chain saturated fatty acid into NA. This review highlights recent advances in the biosynthesis, the sources, and the biotechnological aspects of NA.

A Rapid Method for the Determination of Fucoxanthin in Diatom.

Fucoxanthin is a natural pigment found in microalgae, especially diatoms and Chrysophyta. Recently, it has been shown to have anti-inflammatory, anti-tumor, and anti-obesityactivity in humans. Phaeodactylum tricornutum is a diatom with high economic potential due to its high content of fucoxanthin and eicosapentaenoic acid. In order to improve fucoxanthin production, physical and chemical mutagenesis could be applied to generate mutants. An accurate and rapid method to assess the fucoxanthin content is a prerequisite for a high-throughput screen of mutants. In this work, the content of fucoxanthin in P. tricornutum was determined using spectrophotometry instead of high performance liquid chromatography (HPLC). This spectrophotometric method is easier and faster than liquid chromatography and the standard error was less than 5% when compared to the HPLC results. Also, this method can be applied to other diatoms, with standard errors of 3-14.6%. It provides a high throughput screening method for microalgae strains producing fucoxanthin.

Bioremediation of wastewater from edible oil refinery factory using oleaginous microalga Desmodesmus sp. S1.

Edible oil industry produced massive wastewater, which requires extensive treatment to remove pungent smell, high phosphate, carbon oxygen demand (COD), and metal ions prior to discharge. Traditional anaerobic and aerobic digestion could mainly reduce COD of the wastewater from oil refinery factories (WEORF). In this study, a robust oleaginous microalga Desmodesmus sp. S1 was adapted to grow in WEORF. The biomass and lipid content of Desmodesmus sp. S1 cultivated in the WEORF supplemented with sodium nitrate were 5.62 g·L(-1) and 14.49%, whereas those in the WEORF without adding nitrate were 2.98 g·L(-1) and 21.95%. More than 82% of the COD and 53% of total phosphorous were removed by Desmodesmus sp. S1. In addition, metal ions, including ferric, aluminum, manganese and zinc were also diminished significantly in the WEORF after microalgal growth, and pungent smell vanished as well. In comparison with the cells grown in BG-11 medium, the cilia-like bulges and wrinkles on the cell surface of Desmodesmus sp. S1 grown in WEORF became out of order, and more polyunsaturated fatty acids were detected due to stress derived from the wastewater. The study suggests that growing microalgae in WEORF can be applied for the dual roles of nutrient removal and biofuel feedstock production.