Scholarly Work - Center for Biofilm Engineering
Permanent URI for this collectionhttps://scholarworks.montana.edu/handle/1/9335
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Item In Silico Analysis of Functionalized Hydrocarbon Production Using Ehrlich Pathway and Fatty Acid Derivatives in an Endophytic Fungus(MDPI, 2021-05) Hunt, Kristopher A.; Mallette, Natasha D.; Peyton, Brent M.; Carlson, Ross P.Functionalized hydrocarbons have various ecological and industrial uses, from signaling molecules and antifungal/antibacterial agents to fuels and specialty chemicals. The potential to produce functionalized hydrocarbons using the cellulolytic, endophytic fungus, Ascocoryne sarcoides, was quantified using genome-enabled, stoichiometric modeling. In silico analysis identified available routes to produce these hydrocarbons, including both anabolic- and catabolic-associated strategies, and determined correlations between the type and size of the hydrocarbons and culturing conditions. The analysis quantified the limits of the wild-type metabolic network to produce functionalized hydrocarbons from cellulose-based substrates and identified metabolic engineering targets, including cellobiose phosphorylase (CP) and cytosolic pyruvate dehydrogenase complex (PDHcyt). CP and PDHcyt activity increased the theoretical production limits under anoxic conditions where less energy was extracted from the substrate. The incorporation of both engineering targets resulted in near-complete conservation of substrate electrons in functionalized hydrocarbons. The in silico framework was integrated with in vitro fungal batch growth experiments to support O2 limitation and functionalized hydrocarbon production predictions. The metabolic reconstruction of this endo-phytic filamentous fungus describes pathways for both specific and general production strategies of 161 functionalized hydrocarbons applicable to many eukaryotic hosts.Item Integrating CUREs in Ongoing Research: Undergraduates as Active Participants in the Discovery of Biodegrading Thermophiles(2021-09) Peyton, Brent M.; Skorupa, Dana J.Research-based courses are a powerful way to engage undergraduates in the scientific process while simultaneously teaching participants relevant laboratory, analysis, and scientific communication skills. In most programs, students conduct a simulated project which effectively improves student conceptions of scientific thinking but does not produce research-quality data. The course described here delivered an authentic research experience by assigning undergraduates an objective from an active grant-funded project. Participants contributed to research aimed at culturing biodegrading thermophiles from hot springs in Yellowstone National Park. Students participated in a backcountry field experience, collecting environmental samples of their choosing and determining appropriate culturing conditions. Following high-temperature incubations, 16S rRNA gene sequencing identified enriched microbial populations, with analytical and microscopy methods tracking degradation and growth. Importantly, several teams successfully cultivated thermophilic plastic-degrading consortia. Student learning was assessed using several methods, including grade distributions on assignments and statistical comparisons of pre- and posttests. A consistent and, in most cases, statistically significant increase was observed in the students’ posttest scores. The grade distribution on summative assessments also suggests that students achieved the desired learning outcomes. Student perceptions of their learning and experience gains were high, with participants reporting improvements in components emphasized in the research activities. Overall, the findings highlight how involving undergraduates in real-world research projects can enhance student interest and ownership of scientific research, along with contributing quality data that inform active studies.Item Validating an Automated Nucleic Acid Extraction Device for Omics in Space Using Whole Cell Microbial Reference Standards(2020-08) Urbaniak, Camilla; Wong, Season; Tighe, Scott; Arumugam, Arunkumar; Liu, Bo; Parker, Ceth W.; Wood, Jason M.; Singh, Nitin K.; Skorupa, Dana J.; Peyton, Brent M.; Jenson, Ryan; Karouia, Fathi; Dragon, Julie; Venkateswaran, KasthuriNASA has made great strides in the past five years to develop a suite of instruments for the International Space Station in order to perform molecular biology in space. However, a key piece of equipment that has been lacking is an instrument that can extract nucleic acids from an array of complex human and environmental samples. The Omics in Space team has developed the μTitan (simulated micro(μ) gravity tested instrument for automated nucleic acid) system capable of automated, streamlined, nucleic acid extraction that is adapted for use under microgravity. The μTitan system was validated using a whole cell microbial reference (WCMR) standard comprised of a suspension of nine bacterial strains, titrated to concentrations that would challenge the performance of the instrument, as well as to determine the detection limits for isolating DNA. Quantitative assessment of system performance was measured by comparing instrument input challenge dose vs recovery by Qubit spectrofluorometry, qPCR, Bioanalyzer, and Next Generation Sequencing. Overall, results indicate that the μTitan system performs equal to or greater than a similar commercially available, earth-based, automated nucleic acid extraction device. The μTitan system was also tested in Yellowstone National Park (YNP) with the WCMR, to mimic a remote setting, with limited resources. The performance of the device at YNP was comparable to that in a laboratory setting. Such a portable, field-deployable, nucleic extraction system will be valuable for environmental microbiology, as well as in health care diagnostics.Item Microbial community changes during a toxic cyanobacterial bloom in an alkaline Hungarian lake(2018-08) Bell, Tisza A. S.; Feldoldi, Tamas; Sen-Kilic, Emel; Vasas, Gabor; Fields, Matthew W.; Peyton, Brent M.The Carpathian Basin is a lowland plain located mainly in Hungary. Due to the nature of the bedrock, alluvial deposits, and a bowl shape, many lakes and ponds of the area are characterized by high alkalinity. In this study, we characterized temporal changes in eukaryal and bacterial community dynamics with high throughput sequencing and relate the changes to environmental conditions in Lake Velence located in Fejer county, Hungary. The sampled Lake Velence microbial populations (algal and bacterial) were analyzed to identify potential correlations with other community members and environmental parameters at six timepoints over 6weeks in the Spring of 2012. Correlations between community members suggest a positive relationship between certain algal and bacterial populations (e.g. Chlamydomondaceae with Actinobacteria and Acidobacteria), while other correlations allude to changes in these relationships over time. During the study, high nitrogen availability may have favored non-nitrogen fixing cyanobacteria, such as the toxin-producing Microcystis aeruginosa, and the eutrophic effect may have been exacerbated by high phosphorus availability as well as the high calcium and magnesium content of the Carpathian Basin bedrock, potentially fostering exopolymer production and cell aggregation. Cyanobacterial bloom formation could have a negative environmental impact on other community members and potentially affect overall water quality as well as recreational activities. To our knowledge, this is the first prediction for relationships between photoautotrophic eukaryotes and bacteria from an alkaline, Hungarian lake.Item Assessment of Nannochloropsis gaditana growth and lipid accumulation with increased inorganic carbon delivery(2018-04) Pedersen, Todd C.; Gardner, Robert D.; Peyton, Brent M.Algal biomass refineries for sustainable transportation fuels, in particular biodiesel, will benefit from algal strain enhancements to improve biomass and lipid productivity. Specifically, the supply of inorganic carbon to microalgal cultures represents an area of great interest due to the potential for improved growth of microalgae and the possibility for incorporation with CO2 mitigation processes. Combinations of bicarbonate (HCO3−) salt addition and application of CO2 to control pH have shown compelling increases in growth rate and lipid productivity of fresh water algae. Here, focus was placed on the marine organism, Nannochloropsis gaditana, to investigate growth and lipid accumulation under various strategies of enhanced inorganic carbon supply. Three gas application strategies were investigated: continuous sparging of atmospheric air, continuous sparging of 5% CO2 during light hours until nitrogen depletion, and continuous sparging of atmospheric air supplemented with 5% CO2 for pH control between 8.0 and 8.3. These gas sparging schemes were combined with addition of low concentrations (5 mM) of sodium bicarbonate at inoculation and high concentration (50 mM) of sodium bicarbonate amendments just prior to nitrogen depletion. The optimum scenario observed for growth of N. gaditana under these inorganic carbon conditions was controlling pH with 5% CO2 on demand, which increased both growth rate and lipid accumulation. Fatty acid methyl esters were primarily comprised of C16:0 (palmitic) and C16:1 (palmitoleic) aliphatic chains. Additionally, the use of high concentration (50 mM) of bicarbonate amendments further improved lipid content (up to 48.6%) under nitrogen deplete conditions when paired with pH-controlled strategies.Item Effects of shear stress and substrate loading rate on pseudomonas aeruginosa biofilm thickness and density(1996-01) Peyton, Brent M.Thickness, roughness, and density data were obtained from monopopulation Pseudomonas aeruginosa biofilms grown in an annular reactor under different substrate loading rates and shear stresses. Biofilm thickness showed significant increases, up to approx. 30 μm, with increasing substrate loading rate. Shear stress had no significant effect on thickness. Biofilm roughness, as measured by the standard deviation of the thickness, increased with increasing thickness. Areal mass density significantly increased with substrate loading rate and was not significantly affected by shear stress. Substrate loading and shear stress did not significantly affect volumetric mass density. A numerical expression that describes changes volumetric density as a function of biofilm depth is presented.Item Biofilm surface positioning(1990) Lewandowski, Zbigniew; Walser, Gabriele Sabine; Larsen, R. W.; Peyton, Brent M.; Characklis, William G.Item Interactions between process waters, microbial biofilms, and metal substrata(1990) Characklis, William G.; Lee, Whonchee; Peyton, Brent M.; Lewandowski, ZbigniewItem A statistical analysis of the effect of substrate utilization and shear stress on the kinetics of biofilm detachment(1993-03) Peyton, Brent M.; Characklis, William G.One of the least understood processes affecting biofilm accumulation is detachment. Detachment is the removal of cells and cell products from an established biofilm and subsequent entrainment in the bulk liquid. The goal of this research was to determine the effects of shear stress and substrate loading rate on the rate of biofilm detachment. Monopopulation Pseudomonas aeruginosa and undefined mixed population biofilms were grown on glucose in a RotoTorque biofilm reactor. Three levels of shear stress and substrate loading rate were used to determine their effects on the rate of detachment. Suspended cell concentrations were monitored to determine detachment rates, while other variables were measured to determine their influence on the detachment rate. Results indicate that detachment rate is directly related to biofilm growth rate and that factors which limit growth rate will also limit detachment rate. No significant influence of shear on detachment rate was observed. A new kinetic expression that incorporates substrate utilization rate, yield, and biofilm thickness was compared to published detachment expressions and gives a better correlation of data obtained both in this research and from previous research projects, for both mono- and mixed-population biofilms.Item Fundamentals of Biofilm Processes(1988-08) Characklis, William G.; Larsen, R. W.; Peyton, Brent M.