Potential for Biomass Production and Remediation by Cultivation of the Marine Model Diatom Phaeodactylum tricornutum in Oil Field Produced Wastewater Media

While oilfield produced water (PW) is one of the largest, unclaimed wastewater streams of the oil industry, it could potentially be used as a cultivation medium for microalgae. Microalgae could help with the remediation of this water while also delivering biomass that can be transformed into valuable byproducts such as biofuels. The coupling of these two purposes is expected to cut production costs of biofuels while aiding environmental protection. In this study, we compared the cultivation capacity of the marine model diatom Phaeodactylum tricornutum in media at varying salinities and in media composed of PW from two oilfields in the Central Valley of California that differed drastically in the concentration of inorganic and organic constituents. Specifically, we measured the carrying capacity of these media, the maximum growth rates of P. tricornutum, its cellular lipid accumulation capacity, and its capacity to remediate the most polluted PW source. Our study shows that P. tricornutum can successfully adjust to the tested cultivation media through processes of short-term acclimation and long-term adaptation. Furthermore, the cultivation of P. tricornutum in the most heavily polluted PW source led to significant increases in cell yield and improved photosynthetic capacity during the stationary phase, which could be attributed chiefly to the higher levels of nitrate present in this PW source. Chemical water analyses also demonstrated the capability of P. tricornutum to remediate major nutrient content and potentially harmful elements like fluorine and copper. Because P. tricornutum is amenable to advanced genetic engineering, which could be taken advantage of to improve its cultivation resilience and productivity in an economic setting, we propose this study as a step towards essential follow-up studies that will identify the genetic regulation behind its growth in oilfield PW media and its remediation of the PW constituents

Transcriptional Orchestration of the Global Cellular Response of a Model Pennate Diatom to Diel Light Cycling under Iron Limitation.

Environmental fluctuations affect distribution, growth and abundance of diatoms in nature, with iron (Fe) availability playing a central role. Studies on the response of diatoms to low Fe have either utilized continuous (24 hr) illumination or sampled a single time of day, missing any temporal dynamics. We profiled the physiology, metabolite composition, and global transcripts of the pennate diatom Phaeodactylum tricornutum during steady-state growth at low, intermediate, and high levels of dissolved Fe over light:dark cycles, to better understand fundamental aspects of genetic control of physiological acclimation to growth under Fe-limitation. We greatly expand the catalog of genes involved in the low Fe response, highlighting the importance of intracellular trafficking in Fe-limited diatoms. P. tricornutum exhibited transcriptomic hallmarks of slowed growth leading to prolonged periods of cell division/silica deposition, which could impact biogeochemical carbon sequestration in Fe-limited regions. Light harvesting and ribosome biogenesis transcripts were generally reduced under low Fe while transcript levels for genes putatively involved in the acquisition and recycling of Fe were increased. We also noted shifts in expression towards increased synthesis and catabolism of branched chain amino acids in P. tricornutum grown at low Fe whereas expression of genes involved in central core metabolism were relatively unaffected, indicating that essential cellular function is protected. Beyond the response of P. tricornutum to low Fe, we observed major coordinated shifts in transcript control of primary and intermediate metabolism over light:dark cycles which contribute to a new view of the significance of distinctive diatom pathways, such as mitochondrial glycolysis and the ornithine-urea cycle. This study provides new insight into transcriptional modulation of diatom physiology and metabolism across light:dark cycles in response to Fe availability, providing mechanistic understanding for the ability of diatoms to remain metabolically poised to respond quickly to Fe input and revealing strategies underlying their ecological success

Correction: Transcriptional Orchestration of the Global Cellular Response of a Model Pennate Diatom to Diel Light Cycling under Iron Limitation

<p>Correction: Transcriptional Orchestration of the Global Cellular Response of a Model Pennate Diatom to Diel Light Cycling under Iron Limitation</p

Physiological response of <i>P</i>. <i>tricornutum</i> to low Fe across diel cycles.

<p>Diel patterns in cell division and fluctuations of F_v/F_m in <i>P</i>. <i>tricornutum</i> grown at 20 pM, 40 pM, and 400 pM Fe′ over 12:12 L:D cycles, lights on = 9am, lights off = 9pm. (<b>A</b>) Bar plot of the time point averaged percentage of dividing cells shown with standard deviation across biological replicates. (<b>B</b>) Time point averaged F_v/F_m. Legend is shared for both plots and shaded background indicates dark time points.</p

Diel expression of the N-assimilation pathway.

<p>Diagram shows the N-assimilation pathway and enzyme subcellular localizations with the general pattern of expression of genes indicated. Compartments are labeled, and the chloroplast is shown with four membranes. NRT = nitrate transporter, NR = nitrate reductase, NOD = nitric oxide dioxygenase, FNT = formate-nitrite transporter, Fd-NIR = ferredoxin-dependent nitrite reductase, NIR = NADH-dependent nitrite reductase, GS(II/III) = glutamine synthetase, GOGAT = glutamine oxoglutarate aminotransferase, NMO = nitronate monooxygenase. See <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006490#pgen.1006490.s011" target="_blank">S6 Fig</a> for PIDs and normalized transcript abundance.</p

Physiological response of <i>P</i>. <i>tricornutum</i> to varying Fe and illumination.

<p>Average growth rates (biological duplicate for 20 pM and 40 pM Fe′, biological triplicate for 400 pM Fe′ with standard deviation), average divisions per day, and F_v/F_m.</p

Urea cycle and chloroplast ornithine cycle expression and metabolites.

<p>(<b>A</b>) Pathway diagram shows the localization of reactions in the urea cycle, distributed between the mitochondria and the cytosol, and the chloroplast ornithine cycle [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006490#pgen.1006490.ref116" target="_blank">116</a>,<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006490#pgen.1006490.ref119" target="_blank">119</a>]. Compartments are labeled, and the chloroplast is shown with four membranes. Dashed arrow indicates hypothesized transport. unCPS = carbamoyl-phosphate synthase, OTC = ornithine transcarbamylase, ASuS = arginosuccinate synthase, ASL = arginosuccinate lyase, FUM = fumarase, ARG = arginase, ACOAT = acetylornithine aminotransferase, NAGS = N-acetyl glutamate synthase, GACT = glutamate acetyltranferase, AGK = acetylglutamate kinase, AGPR = acetyl-glutamyl phosphate reductase. (<b>B</b>) Metabolite abundance (replicate averaged) over diel cycles in iron-replete conditions, shown normalized to maximum value within each metabolite. Average transcript abundance (across Fe conditions) for arginase is also plotted (with standard deviation).</p

Transcriptional Orchestration of the Global Cellular Response of a Model Pennate Diatom to Diel Light Cycling under Iron Limitation

<div><p>Environmental fluctuations affect distribution, growth and abundance of diatoms in nature, with iron (Fe) availability playing a central role. Studies on the response of diatoms to low Fe have either utilized continuous (24 hr) illumination or sampled a single time of day, missing any temporal dynamics. We profiled the physiology, metabolite composition, and global transcripts of the pennate diatom <i>Phaeodactylum tricornutum</i> during steady-state growth at low, intermediate, and high levels of dissolved Fe over light:dark cycles, to better understand fundamental aspects of genetic control of physiological acclimation to growth under Fe-limitation. We greatly expand the catalog of genes involved in the low Fe response, highlighting the importance of intracellular trafficking in Fe-limited diatoms. <i>P</i>. <i>tricornutum</i> exhibited transcriptomic hallmarks of slowed growth leading to prolonged periods of cell division/silica deposition, which could impact biogeochemical carbon sequestration in Fe-limited regions. Light harvesting and ribosome biogenesis transcripts were generally reduced under low Fe while transcript levels for genes putatively involved in the acquisition and recycling of Fe were increased. We also noted shifts in expression towards increased synthesis and catabolism of branched chain amino acids in <i>P</i>. <i>tricornutum</i> grown at low Fe whereas expression of genes involved in central core metabolism were relatively unaffected, indicating that essential cellular function is protected. Beyond the response of <i>P</i>. <i>tricornutum</i> to low Fe, we observed major coordinated shifts in transcript control of primary and intermediate metabolism over light:dark cycles which contribute to a new view of the significance of distinctive diatom pathways, such as mitochondrial glycolysis and the ornithine-urea cycle. This study provides new insight into transcriptional modulation of diatom physiology and metabolism across light:dark cycles in response to Fe availability, providing mechanistic understanding for the ability of diatoms to remain metabolically poised to respond quickly to Fe input and revealing strategies underlying their ecological success.</p></div

Differentially expressed gene totals.

<p>Total number of genes determined to be significantly differentially expressed (p < 0.05) by the Skillings-Mack test and the LR test in EdgeR (EdgeR) and by both tests. Total number of genes determined to be differentially expressed only by one method or the other (Total Unique) are also shown. The three most abundant unique response types for each test are shown in bold. Ratios of Low Fe and High Fe genes expressed in the dark relative to light are calculated for each test.</p