Meaningful Work, Career Fit, and Professional Well-Being of Pediatric Academicians in the United States.

OBJECTIVE: Examine associations between time spent in academic activities perceived as meaningful and professional well-being among academic pediatrics faculty. METHODS: The sample comprised 248 full-time pediatric faculty (76% female, 81% white, non-Hispanic, 41% instructor or assistant professor) across the United States who completed an online survey in November 2019. Survey items included sociodemographic and professional characteristics, professional well-being measures (Stanford Professional Fulfillment Index; Maslach Burnout Inventory; Intention to Leave Academic Medicine), perceived meaningfulness of academic activities and assigned time to those activities. We defined global career fit as total percentage time assigned to professional activities considered meaningful by individuals, and activity-specific career fit as percentage time assigned to each meaningful professional activity. RESULTS: As global career fit scores increased, professional fulfillment increased (r = 0.45, P < .001), whereas burnout (r = -0.29, P < .001) and intention to leave (r = -0.22, P < .001) decreased. Regarding activity-specific career fit, for individuals who considered patient care meaningful, as assigned time to patient care increased, professional fulfillment decreased (r = -0.14, P = .048) and burnout (r = 0.16, P = .02) and intention to leave (r = 0.26, P < .001) increased. There was no significant correlation between assigned time for teaching, research, or advocacy and professional well-being. Faculty were less likely to intend to leave academic medicine as assigned time increased for administrative or leadership activities if considered meaningful (r = -0.24, P = .01). CONCLUSIONS: Time assigned to meaningful work activities may relate to professional well-being of academic pediatrics faculty. More time assigned to patient care, despite being meaningful, was associated with poor self-reported professional well-being. Effort allocation among diverse academic activities needs to be optimized to improve faculty well-being.

High ethanol titers from cellulose by using metabolically engineered thermophilic, anaerobic microbes.

This work describes novel genetic tools for use in Clostridium thermocellum that allow creation of unmarked mutations while using a replicating plasmid. The strategy employed counter-selections developed from the native C. thermocellum hpt gene and the Thermoanaerobacterium saccharolyticum tdk gene and was used to delete the genes for both lactate dehydrogenase (Ldh) and phosphotransacetylase (Pta). The Δldh Δpta mutant was evolved for 2,000 h, resulting in a stable strain with 40:1 ethanol selectivity and a 4.2-fold increase in ethanol yield over the wild-type strain. Ethanol production from cellulose was investigated with an engineered coculture of organic acid-deficient engineered strains of both C. thermocellum and T. saccharolyticum. Fermentation of 92 g/liter Avicel by this coculture resulted in 38 g/liter ethanol, with acetic and lactic acids below detection limits, in 146 h. These results demonstrate that ethanol production by thermophilic, cellulolytic microbes is amenable to substantial improvement by metabolic engineering.

Metabolic engineering of a thermophilic bacterium to produce ethanol at high yield.

We report engineering Thermoanaerobacterium saccharolyticum, a thermophilic anaerobic bacterium that ferments xylan and biomass-derived sugars, to produce ethanol at high yield. Knockout of genes involved in organic acid formation (acetate kinase, phosphate acetyltransferase, and L-lactate dehydrogenase) resulted in a strain able to produce ethanol as the only detectable organic product and substantial changes in electron flow relative to the wild type. Ethanol formation in the engineered strain (ALK2) utilizes pyruvate:ferredoxin oxidoreductase with electrons transferred from ferredoxin to NAD(P), a pathway different from that in previously described microbes with a homoethanol fermentation. The homoethanologenic phenotype was stable for >150 generations in continuous culture. The growth rate of strain ALK2 was similar to the wild-type strain, with a reduction in cell yield proportional to the decreased ATP availability resulting from acetate kinase inactivation. Glucose and xylose are co-utilized and utilization of mannose and arabinose commences before glucose and xylose are exhausted. Using strain ALK2 in simultaneous hydrolysis and fermentation experiments at 50 degrees C allows a 2.5-fold reduction in cellulase loading compared with using Saccharomyces cerevisiae at 37 degrees C. The maximum ethanol titer produced by strain ALK2, 37 g/liter, is the highest reported thus far for a thermophilic anaerobe, although further improvements are desired and likely possible. Our results extend the frontier of metabolic engineering in thermophilic hosts, have the potential to significantly lower the cost of cellulosic ethanol production, and support the feasibility of further cost reductions through engineering a diversity of host organisms.

Strain and bioprocess improvement of a thermophilic anaerobe for the production of ethanol from wood.

BACKGROUND: The thermophilic, anaerobic bacterium Thermoanaerobacterium saccharolyticum digests hemicellulose and utilizes the major sugars present in biomass. It was previously engineered to produce ethanol at yields equivalent to yeast. While saccharolytic anaerobes have been long studied as potential biomass-fermenting organisms, development efforts for commercial ethanol production have not been reported. RESULTS: Here, we describe the highest ethanol titers achieved from T. saccharolyticum during a 4-year project to develop it for industrial production of ethanol from pre-treated hardwood at 51-55 °C. We describe organism and bioprocess development efforts undertaken to improve ethanol production. The final strain M2886 was generated by removing genes for exopolysaccharide synthesis, the regulator perR, and re-introduction of phosphotransacetylase and acetate kinase into the methyglyoxal synthase gene. It was also subject to multiple rounds of adaptation and selection, resulting in mutations later identified by resequencing. The highest ethanol titer achieved was 70 g/L in batch culture with a mixture of cellobiose and maltodextrin. In a "mock hydrolysate" Simultaneous Saccharification and Fermentation (SSF) with Sigmacell-20, glucose, xylose, and acetic acid, an ethanol titer of 61 g/L was achieved, at 92 % of theoretical yield. Fungal cellulases were rapidly inactivated under these conditions and had to be supplemented with cellulosomes from C. thermocellum. Ethanol titers of 31 g/L were reached in a 100 L SSF of pre-treated hardwood and 26 g/L in a fermentation of a hardwood hemicellulose extract. CONCLUSIONS: This study demonstrates that thermophilic anaerobes are capable of producing ethanol at high yield and at titers greater than 60 g/L from purified substrates, but additional work is needed to produce the same ethanol titers from pre-treated hardwood.

Anaerobic detoxification of acetic acid in a thermophilic ethanologen.

BACKGROUND: The liberation of acetate from hemicellulose negatively impacts fermentations of cellulosic biomass, limiting the concentrations of substrate that can be effectively processed. Solvent-producing bacteria have the capacity to convert acetate to the less toxic product acetone, but to the best of our knowledge, this trait has not been transferred to an organism that produces ethanol at high yield. RESULTS: We have engineered a five-step metabolic pathway to convert acetic acid to acetone in the thermophilic anaerobe Thermoanaerobacterium saccharolyticum. The first steps of the pathway, a reversible conversion of acetate to acetyl-CoA, are catalyzed by the native T. saccharolyticum enzymes acetate kinase and phosphotransacetylase. ack and pta normally divert 30% of catabolic carbon flux to acetic acid; however, their re-introduction in evolved ethanologen strains resulted in virtually no acetic acid production. Conversion between acetic acid and acetyl-CoA remained active, as evidenced by rapid (13)C label transfer from exogenous acetate to ethanol. Genomic re-sequencing of six independently evolved ethanologen strains showed convergent mutations in the hfs hydrogenase gene cluster, which when transferred to wildtype T. saccharolyticum conferred a low acid production phenotype. Thus, the mutated hfs genes effectively separate acetic acid production and consumption from central metabolism, despite their intersecting at the common intermediate acetyl-CoA. To drive acetic acid conversion to a less inhibitory product, the enzymes thiolase, acetoacetate:acetate CoA-transferase, and acetoacetate decarboxylase were assembled in T. saccharolyticum with genes from thermophilic donor organisms that do not natively produce acetone. The resultant strain converted acetic acid to acetone and ethanol while maintaining a metabolic yield of 0.50 g ethanol per gram carbohydrate. CONCLUSIONS: Conversion of acetic acid to acetone results in improved ethanol productivity and titer and is an attractive low-cost solution to acetic acid inhibition.