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RNA-seq comparison of transcript abundance in a PDC-producing N. aromaticivorans strain (12444PDC) grown in the presence of dehydrodiconiferyl alcohol (DC-A) and glucose, vanillin and glucose, ferulic acid and glucose, or glucose alone
The present disclosure relates to a genetically modified microbial cell that includes a first genetic modification resulting in the expression of an exogenous vanillate demethylase, such that the microbial cell is capable of metabolizing an S-lignin decomposition product and producing 2-pyrone-4,6-dicarboxylate (PDC).
Drilling activities account for 30% to 57% of the cost to develop and install a geothermal plant. Therefore, an accurate representation of the cost to drill a well is paramount in techno-economic analysis to determine the feasibility of a geothermal power project. In 2022, the National Renewable Energy Laboratory (NREL) endeavored to revise the U.S. Department of Energy (DOE) GeoVision baseline drilling cost curves due to extensive improvement in drilling rates at the Utah Frontier Observatory Research in Geothermal Energy (FORGE) demonstration site. That effort did not culminate in the recommendation of new curves because the actual project costs did not match the reported performance improvements and were at or above the GeoVision baseline. The need for another iteration of this analysis has arisen from industry record drilling performance reported by recent commercial field-scale and demonstration projects, including Fervo Energy's Cape Station, the Utah FORGE 16B(78)-32 demonstration and the Geysers Power Company's GDC-36 demonstration. Therefore, in this work, we have estimated the resulting industry average rate of penetration (ROP) and bit life and applied these parameters as inputs to the Well Cost Simplified model used in the GeoVision analysis. The resulting revised cost curves show a significant decline from the GeoVision baseline. For vertical wells, the magnitude of cost decline ranges between 12% and 24% while for deviated wells, cost reductions between 18% and 26% are estimated. The revised cost curves are in good agreement with actual cost data and therefore, quantify the economic impact of the utilization of (and advances in) polycrystalline diamond compact (PDC) bit technology and the application of physics-based methodologies that optimize mechanical specific energy.
As part of a U.S. Department of Energy (DOE) Geothermal Technologies Office-funded initiative, Geysers Power Company, LLC, a subsidiary of Calpine Corporation, has been working to enhance drilling performance at the world’s largest geothermal field, The Geysers, in northern California. In a recent drilling operation of the GDC-36 well, excessive mud losses were encountered, initially addressed through repeated but largely ineffective cement plugging. Ultimately, the most effective strategy was to drill blind through the loss zones, made feasible by the high rate of penetration (ROP) achieved with PDC bits, allowing significant progress before the mud tanks were depleted and water-sensitive argillic formation layers could collapse. In response to these challenges, the project team explored alternative methods to minimize downtime and risks associated with cement plugging and continuous mud loss and to contemplate the driving mechanisms for the losses. Wellbore imaging using Formation MicroImager (FMI) and Ultrasonic Borehole Imager (UBI) tools revealed longitudinal tensile fractures, which were attributed to mud weights exceeding the minimum circumferential stress resulting from the native stress field and formation pressure. This study examines the mud losses encountered and leverages wellbore imaging data to understand the mechanisms behind mud induced tensile fracturing in specific rock facies. Understanding fracture behavior across different lithologies is crucial, as fractures within the reservoir can enhance steam migration throughout the system. The reservoir at The Geysers lies within the Mesozoic Franciscan Assemblage, a tectonic mélange formed by subduction. It consists of metamorphosed turbidite sandstone (greywacke) and mudstone (argillite), oceanic upper crust (including greenstone and chert), and serpentinized ultramafic rocks - each exhibiting distinct geomechanical fracturing properties. The structural fabric of the Franciscan Assemblage was shaped by low-angle Mesozoic thrust faulting and later overprinted by sub-vertical strike-slip structures related to the Pacific-North American plate boundary. A wellbore stability model was developed using core measurements and logs to simulate fracturing scenarios during drilling under varying stress conditions. These simulations guided the development of an optimized mud weight management strategy that should enable adaptive adjustments during drilling, reducing the likelihood of tensile fracturing and mud losses, ultimately improving operational efficiency.
ABSTRACT Carotenoids are lipophilic compounds found in the membranes of various organisms. Individual carotenoids are also commodity chemicals, produced industrially for use as food additives, nutritional supplements, cosmetics, and pharmaceuticals. The alphaproteobacterium Novosphingobium aromaticivorans has previously been established as a potential platform microbe for converting aromatic compounds derived from lignocellulosic plant biomass into valuable extracellular products. Here, we show that N. aromaticivorans DSM 12444 cells naturally produce the carotenoid nostoxanthin, and we construct a set of gene deletion mutants that accumulate β-carotene, lycopene, or zeaxanthin, which are predicted intermediates in nostoxanthin biosynthesis as well as commodity chemicals. We also show that a mutant strain heterologously expressing a CrtW protein accumulates the carotenoid astaxanthin. When grown on vanillate as the carbon source, we find that the levels of carotenoids are not significantly affected by O 2 concentration in the tested range of 5% to 21% O 2 . We also show that these carotenoids are produced at comparable levels when strains are grown in liquor from alkaline pretreated sorghum biomass [sorghum alkaline pretreatment liquor (APL)], which contains a mixture of aromatics. Finally, we construct strains that produce zeaxanthin, β-carotene, or astaxanthin concurrently with 2-pyrone-4,6-dicarboxylic acid, a potential building block for biodegradable polymers, when grown in sorghum APL. Combined, our results show that N. aromaticivorans can simultaneously produce valuable intracellular and extracellular commodities when grown in the presence of either pure aromatics or pretreated lignocellulosic biomass. IMPORTANCE There is economic and environmental interest in generating commodity chemicals from renewable resources, such as lignocellulosic biomass, that can substitute for chemicals derived from fossil fuels. The bacterium Novosphingobium aromaticivorans is a promising microbial platform for producing commodity chemicals from lignocellulosic biomass because it can produce these from compounds in pretreated lignocellulosic biomass, which many industrial microbial catalysts cannot metabolize. Here, we show that N. aromaticivorans can be engineered to produce several valuable carotenoids. We also show that engineered N. aromaticivorans strains can produce these lipophilic chemicals concurrently with the extracellular commodity chemical 2-pyrone-4,6-dicarboxylic acid when grown in a complex liquor obtained from alkaline pretreated lignocellulosic biomass. Concurrent microbial production of valuable intra- and extracellular products can increase the economic value generated from the conversion of lignocellulosic biomass-derived compounds into commodity chemicals and facilitate the separation of water- and membrane-soluble products.
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