Nitrogen-depleted Conditions Research Articles

Valorization of biodiesel-derived crude glycerol for simultaneous biosynthesis of biodegradable polyhydroxybutyrate and exopolysaccharide by the newly isolated Burkholderia sp. SCN-KJ

This study demonstrated that Burkholderia sp. SCN-KJ is a promising novel species for the biovalorization of crude glycerol to polyhydroxybutyrate (PHB) and galactose-rich heteroexopolysaccharide (EPS). Whole-genome and genetic evolution analyses revealed separation of the different clades according to the ANIb and dDDH analyses, which confirmed that Burkholderia sp. SCN-KJ is a novel species. The highest PHB production from crude glycerol was 12.9 ± 0.4 g/L (72.9 ± 2.1 % w/w), with a productivity of 0.46 g/L/h and YP/S of 0.3 g/g at 28 h in a 10 L fermenter. The galactose-rich hetero-EPS began to be produced after nitrogen depletion, resulting in a concentration of 22.4 ± 0.2 g/L at 38 h. Examination of the carbon-to‑nitrogen ratio (C/N) showed that nitrogen-rich condition (C/N 20) was optimal for PHB production, whereas nitrogen-depleted condition promoted EPS production, showing two different extrema. The findings showed that Burkholderia sp. SCN-KJ has the potential to transform the landscape of biovalorization for sustainable production.

Evaluation of Extraction Techniques for Recovery of Microalgal Lipids under Different Growth Conditions.

Microalgal lipids contain a wide array of liposoluble bioactive compounds, but lipid extraction remains a critical limitation for their commercial use. An accelerated solvent extraction (ASE) was used to extract lipids from Chlamydomonas reinhardtii, Arthrospira platensis (Spirulina), and Chlorella vulgaris grown under either standard or nitrogen depletion conditions. Under standard growth conditions, ASE using methanol:chloroform (2:1), methyl tert-butyl ether (MTBE):methanol:water, and ethanol at 100 °C resulted in the highest recovery of total lipids (352 ± 30, 410 ± 32, and 127 ± 15 mg/g biomass from C. reinhardtii, C. vulgaris, and A. platensis, respectively). Similarly, the highest total lipid and triacylglycerols (TAGs) recovery from biomass cultivated under nitrogen depletion conditions was found at 100 °C using methanol:chloroform, for C. reinhardtii (total, 550 ± 21; TAG, 205 ± 2 mg/g biomass) and for C. vulgaris (total, 612 ± 29 mg/g; TAG, 253 ± 7 mg/g biomass). ASE with MTBE:methanol:water at 100 °C yielded similar TAG recovery for C. reinhardtii (159 ± 6 mg/g) and C. vulgaris (200 ± 4 mg/g). Thus, MTBE:methanol:water is suggested as an alternative substitute to replace hazardous solvent mixtures for TAGs extraction with a much lower environmental impact. The extracted microalgal TAGs were rich in palmitic (C16:0), stearic (C18:0), oleic (C18:1,9), linoleic (C18:2n6), and α-linolenic (C18:3n3) acids. Under nitrogen depletion conditions, increased palmitic acid (C16:0) recovery up to 2-fold was recorded from the biomasses of C. reinhardtii and C. vulgaris. This study demonstrates a clear linkage between the extraction conditions applied and total lipid and TAG recovery.

High production of ectoine from methane in genetically engineered Methylomicrobium alcaliphilum 20Z by preventing ectoine degradation

BackgroundMethane is a greenhouse gas with a significant potential to contribute to global warming. The biological conversion of methane to ectoine using methanotrophs represents an environmentally and economically beneficial technology, combining the reduction of methane that would otherwise be combusted and released into the atmosphere with the production of value-added products.ResultsIn this study, high ectoine production was achieved using genetically engineered Methylomicrobium alcaliphilum 20Z, a methanotrophic ectoine-producing bacterium, by knocking out doeA, which encodes a putative ectoine hydrolase, resulting in complete inhibition of ectoine degradation. Ectoine was confirmed to be degraded by doeA to N-α-acetyl-L-2,4-diaminobutyrate under nitrogen depletion conditions. Optimal copper and nitrogen concentrations enhanced biomass and ectoine production, respectively. Under optimal fed-batch fermentation conditions, ectoine production proportionate with biomass production was achieved, resulting in 1.0 g/L of ectoine with 16 g/L of biomass. Upon applying a hyperosmotic shock after high–cell–density culture, 1.5 g/L of ectoine was obtained without further cell growth from methane.ConclusionsThis study suggests the optimization of a method for the high production of ectoine from methane by preventing ectoine degradation. To our knowledge, the final titer of ectoine obtained by M. alcaliphilum 20ZDP3 was the highest in the ectoine production from methane to date. This is the first study to propose ectoine production from methane applying high cell density culture by preventing ectoine degradation.

Identification and characterization of the low molecular mass ferredoxins involved in central metabolism in Heliomicrobium modesticaldum.

The homodimeric Type I reaction center (RC) from Heliomicrobium modesticaldum lacks the PsaC subunit found in Photosystem I and instead uses the interpolypeptide [4Fe-4S] cluster FX as the terminal electron acceptor. Our goal was to identify which of the small mobile dicluster ferredoxins encoded by the H. modesticaldum genome are capable of accepting electrons from the heliobacterial RC (HbRC) and pyruvate:ferredoxin oxidoreductase (PFOR), a key metabolic enzyme. Analysis of the genome revealed seven candidates: HM1_1462 (PshB1), HM1_1461 (PshB2), HM1_2505 (Fdx3), HM1_0869 (FdxB), HM1_1043, HM1_0357, and HM1_2767. Heterologous expression in Escherichia coli and studies using time-resolved optical spectroscopy revealed that only PshB1, PshB2, and Fdx3 are capable of accepting electrons from the HbRC and PFOR. Modeling studies using AlphaFold show that only PshB1, PshB2, and Fdx3 should be capable of docking on PFOR at a positively charged patch that overlays a surface-proximal [4Fe-4S] cluster. Proteomic analysis of wild-type and gene deletion strains ΔpshB1, ΔpshB2, ΔpshB1pshB2, and Δfdx3 grown under nitrogen-replete conditions revealed that Fdx3 is undetectable in the wild-type, ΔpshB1, and Δfdx3 strains, but it is present in the ΔpshB2 and ΔpshB1pshB2 strains, implying that Fdx3 may substitute for PshB2. When grown under nitrogen-deplete conditions, Fdx3 is present in the wild-type and all deletion strains except for Δfdx3. None of the knockout strains demonstrated significant impairment during chemotrophic dark growth on pyruvate, photoheterotrophic light growth on pyruvate, or phototrophic growth on acetate+CO2, indicating a high degree of redundancy among these three electron transfer proteins. Loss of both PshB1 and PshB2, but not FdxB, resulted in poor growth under N2-fixing conditions.

Bacterial endosymbionts of a nitrogen-fixing yeast Rhodotorula mucilaginosa JGTA-S1 - insights into a yet unknown micro-ecosystem.

Rhodotorula mucilaginosa JGTA-S1 is a yeast strain capable of fixing nitrogen and improving nitrogen nutrition in rice plants because of its nitrogen-fixing endobacteria, namely Stutzerimonas (Pseudomonas) stutzeri and Bradyrhizobium sp. To gain a deeper understanding of yeast endosymbionts, we conducted a whole-genome shotgun metagenomic analysis of JGTA-S1 cells grown under conditions of nitrogen sufficiency and deficiency. Our results showed that the endosymbiont population varied depending on the nitrogen regime. Upon mechanical disruption of yeast cells, we obtained endosymbionts in culturable form viz. Bacillus velezensis and Staphylococcus sp. under nitrogen-replete conditions and Lysinibacillus telephonicus., Brevibacillus sp., and Niallia circulans under nitrogen-depleted conditions. S. stutzeri and Bradyrhizobium sp. the previously reported endosymbionts remained unculturable. The culturable endosymbionts Staphylococcus sp. and Bacillus velezensis appear to possess genes for dissimilatory nitrate reduction (DNRA), an alternative pathway for ammonia synthesis. However, our findings suggest that these endosymbionts are facultative as they survive outside the host. The fitness of the yeast was not affected by curing of these microbes. Curing the yeast diazotrophic endosymbionts took a toll on its fitness. Our results also showed that the populations of S. stutzeri and B. velezensis increased significantly under nitrogen-depleted conditions compared to nitrogen-sufficient conditions. The importance of DNRA and nitrogen fixation is also reflected in the metagenomic reads of JGTA-S1.

Production of xylose polysaccharides using newly isolated Chlamydomonas sp. Xylok with nitrogen depletion

The growing interest in sustainable production of polysaccharides derived from microalgae has stimulated our pursuit of a novel biorefinery approach capable of simultaneously generating exopolysaccharide (EPS) and biomass using a new microalgal strain. In the present study, a novel strain, designated as Chlamydomonas sp. Xylok, was successfully isolated from river water contaminated with copper and zinc. This unique strain exhibited remarkable attributes, demonstrating both a high EPS generation capacity and exceptional biomass production compared to previously documented microalgal types. To optimize EPS produced from Chlamydomonas sp. Xylok, a comprehensive investigation of influential parameters, including light intensity, temperature, and crucial nutrient elements, such as nitrogen, phosphorus, calcium, and magnesium, was conducted. Notably, under nitrogen-depleted conditions, the strain exhibited a rapid and remarkable 49 % increase in EPS production within a mere 24-h timeframe compared to normal growth conditions. In addition to its EPS production potential, Chlamydomonas sp. Xylok holds industrial importance owing to its higher yield of xylose compared to other existing microalgal strains. This unique characteristic presents a cost-effective and efficient method for producing xylose, making it more appealing for various commercial applications.

Effects of Four Organic Carbon Sources on the Growth and Astaxanthin Accumulation of Haematococcus lacustris.

The microalga Haematococcus lacustris has a complex life cycle and a slow growth rate, hampering its mass cultivation. Culture of microalgae with organic carbon sources can increase the growth rate. Few studies have evaluated the effects of organic carbon sources on H. lacustris. We compared the vegetative and inductive stages of H. lacustris under autotrophic and mixotrophic conditions using four organic carbon sources: sodium acetate, glycerol, sodium gluconate, and ribose, each at various concentrations (0.325, 0.65, 1.3, and 2.6 g/L). The cell density was increased by 1.3 g/L of glycerol in the vegetative stage. The rapid transition to the inductive stage under nitrogen-depletion conditions caused by 1.3 or 2.6 g/L sodium acetate promoted the accumulation of astaxanthin. The production of astaxanthin by H. lacustris in mass culture using organic carbon sources could increase profitability.

Two plastidial lysophosphatidic acid acyltransferases differentially mediate the biosynthesis of membrane lipids and triacylglycerols in Phaeodactylum tricornutum.

Lysophosphatidic acid acyltransferases (LPAATs) catalyze the formation of phosphatidic acid (PA), a central metabolite in both prokaryotic and eukaryotic organisms for glycerolipid biosynthesis. Phaeodactylum tricornutum contains at least two plastid-localized LPAATs (ptATS2a and ptATS2b), but their roles in lipid synthesis remain unknown. Both ptATS2a and ptATS2b could complement the high temperature sensitivity of the bacterial plsC mutant deficient in LPAAT. In vitro enzyme assays showed that they prefer lysophosphatidic acid over other lysophospholipids. ptATS2a is localized in the plastid inner envelope membrane and CRISPR/Cas9-generated ptATS2a mutants showed compromised cell growth, significantly changed plastid and extra-plastidial membrane lipids at nitrogen-replete condition and reduced triacylglycerols (TAGs) under nitrogen-depleted condition. ptATS2b is localized in thylakoid membranes and its knockout led to reduced growth rate and TAG content but slightly altered molecular composition of membrane lipids. The changes in glycerolipid profiles are consistent with the role of both LPAATs in the sn-2 acylation of sn-1-acyl-glycerol-3-phosphate substrates harboring 20:5 at the sn-1 position. Our findings suggest that both LPAATs are important for membrane lipids and TAG biosynthesis in P. tricornutum and further highlight that 20:5-Lyso-PA is likely involved in the massive import of 20:5 back to the plastid to feed plastid glycerolipid syntheses.

Raman spectral analysis of microbial pigment compositions in vegetative cells and heterocysts of multicellular cyanobacterium

The one-dimensional multicellular cyanobacterium, Anabaena sp. PCC 7120, exhibits a simple topology consisting of two types of cells under the nitrogen-depleted conditions. Although the differentiated (heterocyst) and undifferentiated cells (vegetative cells) were distinguished by their cellular shapes, we found that their internal states, that is, microbial pigment compositions, were distinguished by using a Raman microscope. Almost of Raman bands of the cellular components were assigned to vibrations of the pigments; chlorophyll a, β-carotene, phycocyanin, and allophycocyanin. We found that the Raman spectral measurement can detect the decomposition of both phycocyanin and allophycocyanin, which are components of the light-harvesting phycobilisome complex in the photosystem II. We observed that the Raman bands of phycocyanin and allophycocyanin exhibited more remarkable decrease in the heterocysts when compared to those of chlorophyll a and β-carotene. This result indicated the prior decomposition of phycobilisome in the heterocysts. We show that the Raman measurement is useful to detect the change of pigment composition in the cell differentiation.

NAD+ Synthetase is Required for Free-living and Symbiotic Nitrogen Fixation in the Actinobacterium Frankia casuarinae.

Frankia spp. are multicellular actinobacteria that fix atmospheric dinitrogen (N2) not only in the free-living state, but also in root-nodule symbioses with more than 200 plant species, called actinorhizal plants. To identify novel Frankia genes involved in N2 fixation, we previously isolated mutants of Frankia casuarinae that cannot fix N2. One of these genes, mutant N3H4, did not induce nodulation when inoculated into the host plant Casuarina glauca. Cell lineages that regained the ability to fix N2 as free-living cells were isolated from the mutant cell population. These restored strains also regained the ability to stimulate nodulation. A comparative ana-lysis of the genomes of mutant N3H4 and restored strains revealed that the mutant carried a mutation (Thr584Ile) in the glutamine-dependent NAD+ synthetase gene (Francci3_3146), while restored strains carried an additional suppressor mutation (Asp478Asn) in the same gene. Under nitrogen-depleted conditions, the concentration of NAD(H) was markedly lower in the mutant strain than in the wild type, whereas it was higher in restored strains. These results indicate that glutamine-dependent NAD+ synthetase plays critical roles in both free-living and symbiotic N2 fixation in Frankia.

Discovery of a LuxR-type regulator involved in isoniazid-dependent gene regulation in Mycobacterium smegmatis

ObjectiveMost non-tuberculous mycobacteria exhibit intrinsic resistance against the anti-tuberculosis drug isoniazid (INH). We previously found that a pyrazinamidase/nicotinamidase of Mycobacterium smegmatis, named PzaA, has an enzymatic activity to hydrolyze INH, which may contribute to intrinsic resistance. Furthermore, PzaA expression is strongly induced by INH under nitrogen-depleted conditions, although the precise mechanism of this phenomenon remains unclear. Here, we aimed to reveal the mechanism underlying the INH-dependent induction of PzaA using a transcriptomic approach. MethodsRNA sequencing was performed to identify INH-inducible genes other than pzaA. 5′ rapid amplification of cDNA ends analysis was employed to identify the transcription start sites of INH-induced transcription units. The function of a LuxR-like regulator gene (MSMEI_1050) found within the gene cluster containing pzaA was confirmed by gene deletion and complementation experiments involving INH hydrolysis assay and quantitative reverse transcription PCR. ResultsRNA sequencing revealed 23 genes that INH strongly induced under conditions of nitrogen depletion, 17 of which were in a gene cluster containing pzaA. This cluster comprised at least three transcription units, including a non–INH–inducible monocistronic unit containing MSMEI_1050. Deletion of this gene deprived M. smegmatis of the ability to respond to INH, and complementation restored this ability. ConclusionsMSMEI_1050 plays a key role in INH-dependent gene regulation. The precise mechanism of action is to be determined in future studies.

Ambiguous controls on simulated diazotrophs in the world oceans

Nitrogen fixers, or diazotrophs, play a key role in the nitrogen and carbon cycle of the world oceans. Diazotrophs are capable of utilising atmospheric dinitrogen which is a competitive advantage over generally faster growing ordinary phytoplankton in nitrogen-depleted conditions in the sun-lit surface ocean. In this study we argue that additional competitive advantages must be at play in order to explain the dynamics and distribution of diazotrophs in the global oceans. Backed by growing published evidence we test the effects of preferential grazing (where zooplankton partly avoids diazotrophs) and high-affinity diazotrophic phosphorus uptake in an Earth System Model of intermediate complexity. Our results illustrate that these fundamentally different model assumptions result in a very similar match to observation-based estimates of nitrogen fixation while, at the same time, they imply very different trajectories into our warming future. The latter applies to biomass, fixation rates as well as to the ratio of the two. We conclude that a more comprehensive understanding of the competition between ordinary and diazotrophic phytoplankton will reduce uncertainties in model-based projections of the oceanic N cycle.

Elucidation of Functional Genes Associated with Long Chain-Polyunsaturated Fatty Acids (LC-PUFAs) Metabolism in Oleaginous Diatom Phaeodactylum tricornutum

Phaeodactylum tricornutum is a marine diatom rich in omega-3 fatty acids, a nutraceutical-relevant product. Long chain-polyunsaturated fatty acids (LC-PUFAs) are crucial dietary components for human development and growth. With the availability of genome information and genetic engineering tools, the productivities of OMEGAs have improved, but the functional and organizational relationship of such protein-encoding genes associated with LC-PUFAs biosynthesis is still not clear. Henceforth, our study highlights the conservation pattern, functionality and interaction of LC-PUFAs protein-encoding genes via in silico analysis. The transcriptome and quantitative PCR analysis demonstrates downregulation of ACS4, ELO6b, PTD5a, and MYB106 genes associated with LC-PUFAs synthesis and upregulation of ECoAH and ACAT1 genes associated with β-oxidation in nitrogen-depleted conditions in P. tricornutum. Phylogenomics studies of LC-PUFAs protein-encoding genes show a highly conserved evolutionary pattern in various microalgal lineages. Further, for elucidating the interaction of LC-PUFA metabolic genes, subcellular networks were predicted and pathway enrichment analysis was performed, providing new insights on the crosstalk between LC-PUFA protein-encoding genes (ELO6, PTD5, ACS, and ACL1), regulatory elements (LEC2, MYB, WIN) and transporters (ABCD1). In conclusion, such extensive functional enrichment analysis will undoubtedly aid in the development of genetically engineered algal strains with enhanced production of biomolecules i.e., LC-PUFAs.

Unmasking of CgYor1-Dependent Azole Resistance Mediated by Target of Rapamycin (TOR) and Calcineurin Signaling in Candida glabrata.

ABSTRACTIn this study, 18 predicted membrane-localized ABC transporters of Candida glabrata were deleted individually to create a minilibrary of knockouts (KO). The transporter KOs were analyzed for their susceptibility toward antimycotic drugs. Although CgYOR1 has previously been reported to be upregulated in various azole-resistant clinical isolates of C. glabrata, deletion of this gene did not change the susceptibility to any of the tested azoles. Additionally, Cgyor1Δ showed no change in susceptibility toward oligomycin, which is otherwise a well-known substrate of Yor1 in other yeasts. The role of CgYor1 in azole susceptibility only became evident when the major transporter CgCDR1 gene was deleted. However, under nitrogen-depleted conditions, Cgyor1Δ demonstrated an azole-susceptible phenotype, independent of CgCdr1. Notably, Cgyor1Δ cells also showed increased susceptibility to target of rapamycin (TOR) and calcineurin inhibitors. Moreover, increased phytoceramide levels in Cgyor1Δ and the deletions of regulators downstream of TOR and the calcineurin signaling cascade (Cgypk1Δ, Cgypk2Δ, Cgckb1Δ, and Cgckb2Δ) in the Cgyor1Δ background and their associated fluconazole (FLC) susceptibility phenotypes confirmed their involvement. Collectively, our findings show that TOR and calcineurin signaling govern CgYor1-mediated azole susceptibility in C. glabrata.

Whole set of constitutive promoters for RpoN sigma factor and the regulatory role of its enhancer protein NtrC in Escherichia coli K-12.

The promoter selectivity of Escherichia coli RNA polymerase (RNAP) is determined by its promoter-recognition sigma subunit. The model prokaryote E. coli K-12 contains seven species of the sigma subunit, each recognizing a specific set of promoters. Using genomic SELEX (gSELEX) screening in vitro, we identified the whole set of ‘constitutive’ promoters recognized by the reconstituted RNAP holoenzyme alone, containing RpoD (σ70), RpoS (σ38), RpoH (σ32), RpoF (σ28) or RpoE (σ24), in the absence of other supporting regulatory factors. In contrast, RpoN sigma (σ54), involved in expression of nitrogen-related genes and also other cellular functions, requires an enhancer (or activator) protein, such as NtrC, for transcription initiation. In this study, a series of gSELEX screenings were performed to search for promoters recognized by the RpoN RNAP holoenzyme in the presence and absence of the major nitrogen response enhancer NtrC, the best-characterized enhancer. Based on the RpoN holoenzyme-binding sites, a total of 44 to 61 putative promoters were identified, which were recognized by the RpoN holoenzyme alone. In the presence of the enhancer NtrC, the recognition target increased to 61–81 promoters. Consensus sequences of promoters recognized by RpoN holoenzyme in the absence and presence of NtrC were determined. The promoter activity of a set of NtrC-dependent and -independent RpoN promoters was verified in vivo under nitrogen starvation, in the presence and absence of RpoN and/or NtrC. The promoter activity of some RpoN-recognized promoters increased in the absence of RpoN or NtrC, supporting the concept that the promoter-bound NtrC-enhanced RpoN holoenzyme functions as a repressor against RpoD holoenzyme. Based on our findings, we propose a model in which the RpoN holoenzyme fulfils the dual role of repressor and transcriptase for the same set of genes. We also propose that the promoter recognized by RpoN holoenzyme in the absence of enhancers is the ‘repressive’ promoter. The presence of high-level RpoN sigma in growing E. coli K-12 in rich medium may be related to the repression role of a set of genes needed for the utilization of ammonia as a nitrogen source in poor media. The list of newly identified regulatory targets of RpoN provides insight into E. coli survival under nitrogen-depleted conditions in nature.

In situ monitoring of chlorophyll a fluorescence in Nannochloropsis oceanica cultures to assess photochemical changes and the onset of lipid accumulation during nitrogen deprivation.

In situ chlorophyll a fluorescence measurements were applied to monitor changes in the photochemical variables of Nannochloropsis oceanica cultures under nitrogen-deplete and nitrogen-replete (control) conditions. In addition, growth, lipid, fatty acid, and pigment contents were also followed. In the control culture, growth was promoted along with pigment content, electron transport rate (ETR), and polyunsaturated fatty acids, while total lipid content and fatty acid saturation level diminished. Under nitrogen-deplete conditions, the culture showed a higher de-epoxidation state of the xanthophyll cycle pigments. Fast transients revealed a poor processing efficiency for electron transfer beyond QA , which was in line with the low ETR due to nitrogen depletion. Lipid content and the de-epoxidation state were the first biochemical variables triggered by the change in nutrient status, which coincided with a 20% drop in the in situ effective quantum yield of PSII (ΔF'/Fm '), and a raise in the Vj measurements. A good correlation was found between the changes in ΔF'/Fm ' and lipid content (r = -0.96, p < 0.01). The results confirm the reliability and applicability of in situ fluorescence measurements to monitor lipid induction in N. oceanica.

Enhanced Autophagic Activity Improved the Root Growth and Nitrogen Utilization Ability of Apple Plants under Nitrogen Starvation.

Autophagy is a conserved degradation pathway for recycling damaged organelles and aberrant proteins, and its important roles in plant adaptation to nutrient starvation have been generally reported. Previous studies found that overexpression of autophagy-related (ATG) gene MdATG10 enhanced the autophagic activity in apple roots and promoted their salt tolerance. The MdATG10 expression was induced by nitrogen depletion condition in both leaves and roots of apple plants. This study aimed to investigate the differences in the growth and physiological status between wild type and MdATG10-overexpressing apple plants in response to nitrogen starvation. A hydroponic system containing different nitrogen levels was used. The study found that the reduction in growth and nitrogen concentrations in different tissues caused by nitrogen starvation was relieved by MdATG10 overexpression. Further studies demonstrated the increased root growth and the higher nitrogen absorption and assimilation ability of transgenic plants. These characteristics contributed to the increased uptake of limited nitrogen nutrients by transgenic plants, which also reduced the starvation damage to the chloroplasts. Therefore, the MdATG10-overexpressing apple plants could maintain higher photosynthetic ability and possess better growth under nitrogen starvation stress.

Diazotrophic Cyanobacteria are Associated With a Low Nitrate Resupply to Surface Waters in Lake Tanganyika

In Lake Tanganyika, blooms of nitrogen-fixing (diazotrophic) cyanobacteria emerge, when the upper water column re-stratifies after a period of upwelling and convective mixing. During this seasonal transition, diazotrophic cyanobacteria exploit the abundant phosphate and fix nitrogen after other phytoplankton taxa have consumed the available nitrate. However, it remains less clear, which mechanisms favour diazotrophic cyanobacteria under more heavily stratified conditions with lower levels of excess phosphate and persistent nitrate-depletion. Here, we collected profiles of physicochemical parameters, nutrients and photo-pigments, as well as the medium- to large-sized phytoplankton community during two lake-wide cruises to elucidate to what extent the abundance of diazotrophic cyanobacteria in Lake Tanganyika may be controlled by the nitrate resupply through the thermocline into the euphotic zone. At stations where nitrate was depleted, but phosphate remained available near the surface, high densities of diazotrophic cyanobacteria were associated with a low nitrate supply to surface waters. Our data provide first support for two conceptual scenarios, where the relative position of the thermocline and the euphotic depth may create a functional niche for diazotrophic cyanobacteria: when the upward transport of nitrate into the euphotic zone is reduced by a subjacent thermocline, diazotrophic cyanobacteria, comprising Dolichospermum and Anabaenopsis, are key players in the medium-to large-sized phytoplankton community. By contrast, a thermocline located within the euphotic zone allows for a rapid vertical transport of nitrate for a thriving nitrate-assimilating phytoplankton community that evidently outcompetes diazotrophic cyanobacteria. This study highlights that, under nitrogen-depleted conditions, diazotrophic cyanobacteria can also grow in response to a reduced nutrient resupply to the productive surface waters.

Mitochondrial dynamics and degradation in the oleaginous yeast Lipomyces starkeyi.

Emerging evidence implicates the vital role of mitochondria in lipid consumption and storage, highlighting the intimate link between energy production and saving. Although formation of giant lipid droplets, which is the key hallmark of the oleaginous yeast Lipomyces starkeyi, appears to be regulated in response to changes in mitochondrial shape and metabolism, technical limitations of genetic manipulation have become an obstacle to uncover the mitochondrial behavior in this nonconventional yeast. Here, we established an L. starkeyi strain stably expressing a fluorescent marker for monitoring mitochondrial morphology and degradation and found that mitochondria are mostly fragmented in L. starkeyi cells under fermentable, nonfermentable, and nitrogen depletion conditions. Notably, a fraction of mitochondria-specific fluorescent signals was localized to the vacuole, a lytic organelle in yeast, indicating degradation of mitochondria in those cells. This possible catabolic event was more predominant in cells under nutrient-poor conditions than that in cells under nutrient-rich conditions, concomitantly with lipid droplet formation. Collectively, our studies provide a new tool to investigate mitochondrial dynamics in L. starkeyi and decipher the potential role of mitochondrial degradation in lipid metabolism.

A New Species of Freshwater Algae Nephrochlamys yushanlensis sp. nov. (Selenastraceae, Sphaeropleales) and Its Lipid Accumulation during Nitrogen and Phosphorus Starvation

The new species Nephrochlamys yushanlensis sp. nov. is described from a freshwater plankton sample. A comparison of morphology, 18S rDNA gene and ITS2 sequences, and fatty acid profiles showed that the novel strain represents a new lineage within the genus Nephrochlamys. For the first time with a member of the Selenastraceae, experiments with phosphate and nitrate deprivation were conducted to evaluate changes in biomass, lipid and triacylglycerol (TAGs) accumulation, and composition of fatty acids. Biomass dry weight under simultaneous nitrogen and phosphorus depletion was 1.73g·L-1 , which is significantly lower than the 2.41g·L-1 observed in the control. All conditions of nutrient restriction significantly increased the lipid content in comparison with the control. The largest increase in the total lipid content, reaching 58.64% DW per cell at the end of cultivation, occurred with nitrogen deficiency. Significant increases in TAGs content (to 23.69% and 21.74%, respectively) occurred in phosphorus- and nitrogen-depleted conditions in comparison to the control (16.90%). Oleic (49.8-64.1%), palmitic (21.1-22.7%), and linoleic (8.6-10.3%) acids were the dominant fatty acids when cultured on standard BBM medium, as well as with the shortage of nutrients. Phosphorus deprivation as well as absence of both nitrogen and phosphorus led to the appearance of FAMEs α-linolenic (1.5-4.1%) and stearidonic (1.0-1.8%) acids. In general, FAME profiles revealed that the relative percentage of saturated and monounsaturated fatty acids increased (88.9% of total fatty acids) in nitrogen-depletion conditions, suggesting this strain may be suitable for biodiesel production.