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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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The potential of caproate (hexanoate) production using Clostridium kluyveri syntrophic cocultures with Clostridium acetobutylicum or Clostridium saccharolyticum

Caproate (hexanoate) and other medium-chain fatty acids are valuable platform chemicals produced by processes utilizing petroleum or plant oil. Clostridium kluyveri , growing on short chain alcohols (notably ethanol) and carboxylic acids (such as acetate) is noted for its ability to perform chain elongation to produce 4- to 8-carbon carboxylates. C. kluyveri has been studied in monoculture and coculture conditions, which lead to relatively modest carboxylate titers after long fermentation times. To assess the biosynthetic potential of C. kluyveri for caproate production from sugars through coculture fermentations, in the absence of monoculture data in the literature suitable for our coculture experiments, we first explored C. kluyveri monocultures. Some monocultures achieved caproate titers of 150 to over 200 mM in 40–50 h with a production rate of 7.9 mM/h. Based on that data, we then explored two novel, syntrophic coculture partners for producing caproate from sugars: Clostridium acetobutylicum and Clostridium saccharolyticum . Neither species has been cocultured with C. kluyveri before, and both demonstrate promising results. Our experiments of C. kluyveri monocultures and C. kluyveri — C. saccharolyticum cocultures demonstrate exceptionally high caproate titers (145–200 mM), fast production rates (3.25–8.1 mM/h), and short fermentation times (18–45 h). These results represent the most caproate produced by a C. kluyveri coculture in the shortest known fermentation time. We also explored the possibility of heterologous cell fusion between the coculture pairs similar to the results seen previously in our group with C. acetobutylicum and Clostridium ljungdahlii. Fusion events were observed only in the C. acetobutylicum — C. kluyveri coculture pair, and we offer an explanation for the lack of fusion between C. saccharolyticum and C. kluyveri . This work supports the promise of coculture biotechnology for sustainable production of caproate and other platform chemicals.

59 BASIC BIOLOGICAL SCIENCES↗

Short Chain Fatty Acids and Bacterial Taxa Associated with Reduced Salmonella enterica serovar I 4,[5],12:i:- Shedding in Swine Fed a Diet Supplemented with Resistant Potato Starch

Salmonella enterica serovar I 4,[5],12:i:- is a foodborne pathogen of concern because many isolates are multidrug-resistant (resistant to ≥3 antimicrobial classes) and metal tolerant. In this study, three in-feed additives were individually tested for their ability to reduce Salmonella I 4,[5],12:i:- shedding in swine: resistant potato starch (RPS), high amylose corn starch, and a fatty acid blend, compared with a standard control diet over 21 days. Only RPS-fed pigs exhibited a reduction in Salmonella fecal shedding, different bacterial community compositions, and different cecal short chain fatty acid (SCFA) profiles relative to control animals. Within the RPS treatment group, pigs shedding the least Salmonella tended to have greater cecal concentrations of butyrate, valerate, caproate, and succinate. Additionally, among RPS-fed pigs, several bacterial taxa (Prevotella_7, Olsenella, and Bifidobacterium, and others) exhibited negative relationships between their abundances of and the amount of Salmonella in the feces of their hosts. Many of these same taxa also had significant positive associations with cecal concentrations of butyrate, valerate, caproate, even though they are not known to produce these SCFAs. Together, these data suggest the RPS-associated reduction in Salmonella shedding may be dependent on the establishment of bacterial cross feeding interactions that result in the production of certain SCFAs. However, directly feeding a fatty acid mix did not replicate the effect. RPS supplementation could be an effective means to reduce multidrug-resistant (MDR) S. enterica serovar I 4,[5],12:i:- in swine, provided appropriate bacterial communities are present in the gut.

59 BASIC BIOLOGICAL SCIENCES↗

Engineering the cellulolytic extreme thermophile Caldicellulosiruptor bescii to reduce carboxylic acids to alcohols using plant biomass as the energy source

Abstract Caldicellulosiruptor bescii is the most thermophilic cellulolytic organism yet identified (Topt 78 °C). It grows on untreated plant biomass and has an established genetic system thereby making it a promising microbial platform for lignocellulose conversion to bio-products. Here, we investigated the ability of engineered C. bescii to generate alcohols from carboxylic acids. Expression of aldehyde ferredoxin oxidoreductase (aor from Pyrococcus furiosus) and alcohol dehydrogenase (adhA from Thermoanaerobacter sp. X514) enabled C. bescii to generate ethanol from crystalline cellulose and from biomass by reducing the acetate produced by fermentation. Deletion of lactate dehydrogenase in a strain expressing the AOR–Adh pathway increased ethanol production. Engineered strains also converted exogenously supplied organic acids (isobutyrate and n-caproate) to the corresponding alcohol (isobutanol and hexanol) using both crystalline cellulose and switchgrass as sources of reductant for alcohol production. This is the first instance of an acid to alcohol conversion pathway in a cellulolytic microbe.

Biotechnology & Applied Microbiology↗

An approach towards plastic scintillators from thermally activated delayed fluorescent dyes and cross-linkable bismuth compounds

Cross-linked polyvinyltoluene (PVT) based plastic scintillators loaded with dyes exhibiting thermally activated delayed fluorescence (TADF) and various amounts of (caproate)di-(methacrylate) bismuth (CMB) were developed. The co-polymerization of vinyl toluene monomer loaded with different weight ratios of CMB and 1 wt% of TADF dye yielded highly transparent cross-linked plastic scintillators with intense green photoluminescence. The thermal and mechanical properties of the scintillators were investigated using thermogravimetric analysis (TGA), differential scanning calorimetry (DSC) and micro indentation techniques. Here, their performance for radiation detection was assessed by performing gamma spectroscopy using a Cs-137 source and benchmarked to commercial plastic scintillators. The loading of CMB was guided by Monte Carlo N-Particle (MCNP) simulations. The results of this study show that the cross-linking approach and the use of TADF dyes yield scintillators with good mechanical properties with a uniaxial yield strength up to ~66 MPa for compositions loaded with 40 wt% of CMB and high optical quality. However, the light yield in gamma spectroscopy experiments is found to be reduced with the addition of CMB, suggesting a trade-off between mechanical yield strength and light yield in gamma spectroscopy.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Supplemental selenium source on gut health: insights on fecal microbiome and fermentation products of growing puppies

ABSTRACT Selenium is an essential trace element that can modulate the gut microbiome with an impact on host health. The present study aimed to evaluate the effects of organic (selenium-enriched yeast) vs inorganic (sodium selenite) selenium source on fecal end-fermentation products and gut microbiome of puppies from 20 to 52 weeks of age. Alpha and beta diversity of the gut bacterial community were affected by age but not by gender or selenium source. The relative abundance of taxa was differently affected by age, and the DNA concentration of all selected bacterial groups increased with age, although total volatile fatty acids (VFA), acetate, propionate, caproate and lactate concentrations decreased. Organic selenium was associated with a higher concentration of total VFA, propionate and butyrate, a higher number of DNA copies of Lactobacillus, and a trend to lower DNA copies of Escherichia coli. Effects on fecal microbiome during growth differed with selenium source. Females had higher fecal end-fermentation products related to protein degradation, whereas males had higher DNA concentration of Bifidobacterium. Organic selenium might be beneficial over inorganic for dog food supplementation due to the positive modulation of the gut microbiome observed in puppies.

Pereira, Ana Margarida↗

Establishing Butyribacterium methylotrophicum as a Platform Organism for the Production of Biocommodities from Liquid C 1 Metabolites

Using the Wood-Ljungdahl pathway, acetogens can nonphotosynthetically fix gaseous C 1 molecules, preventing them from entering the atmosphere. Many acetogens can also grow on liquid C 1 compounds such as formate and methanol, which avoid the storage and mass transfer issues associated with gaseous C 1 compounds. Substrate redox state also plays an important role in acetogen metabolism and can modulate products formed by these organisms. Butyribacterium methylotrophicum is an acetogen known for its ability to synthesize longer-chained molecules such as butyrate and butanol, which have significantly higher values than acetate or ethanol, from one-carbon (C 1 ) compounds. We explored B. methylotrophicum’s C 1 metabolism by varying substrates, substrate concentrations, and substrate feeding strategies to improve four-carbon product titers. Our results showed that formate utilization by B. methylotrophicum favored acetate production and methanol utilization favored butyrate production. Cofeeding of both substrates produced a high butyrate titer of 4 g/liter when methanol was supplied in excess to formate. Testing of formate feeding strategies, in the presence of methanol, led to further increases in the butyrate to acetate ratio. Mixotrophic growth of liquid and gaseous C 1 substrates expanded the B. methylotrophicum product profile, as ethanol, butanol, and lactate were produced under these conditions. We also showed that B. methylotrophicum is capable of producing caproate, a six-carbon product, presumably through chain elongation cycles of the reverse β-oxidation pathway. Furthermore, we demonstrated butanol production via heterologous gene expression. Finally, our results indicate that both selection of appropriate substrates and genetic engineering play important roles in determining titers of desired products.

09 BIOMASS FUELS↗

Syntrophus aciditrophicus sp. nov., a new anaerobic bacterium that degrades fatty acids and benzoate in syntrophic association with hydrogen-using microorganisms

Strain SBT is a new, strictly anaerobic, gram-negative, nonmotile, non-sporeforming, rod-shaped bacterium that degrades benzoate and certain fatty acids in syntrophic association with hydrogen/formate-using microorganisms. Strain SBT produced approximately 3 mol of acetate and 0.6 mol of methane per mol of benzoate in coculture with Methanospirillum hungatei strain JF1. Saturated fatty acids, some unsaturated fatty acids, and methyl esters of butyrate and hexanoate also supported growth of strain SBT in coculture with Desulfovibrio strain G11. Strain SBT grew in pure culture with crotonate, producing acetate, butyrate, caproate, and hydrogen. The molar growth yield was 17 +/- 1 g cell dry mass per mol of crotonate. Strain SBT did not grow with fumarate, iron(III), polysulfide, or oxyanions of sulfur or nitrogen as electron acceptors with benzoate as the electron donor. The DNA base composition of strain SBT was 43.1 mol% G+C. Analysis of the 16 S rRNA gene sequence placed strain SBT in the delta-subdivision of the Proteobacteria, with sulfate-reducing bacteria. Strain SBT was most closely related to members of the genus Syntrophus. The clear phenotypic and genotypic differences between strain SBT and the two described species in the genus Syntrophus justify the formation of a new species, Syntrophus aciditrophicus.

Non-NASA Center↗