Microbiology Commons^™

Jmh Dissertation 2016.Pdf, Jennifer Hayashi

Jennifer Hayashi

Transcriptomic And Proteomic Dynamics In The Metabolism Of A Diazotrophic Cyanobacterium, Cyanothece Sp. Pcc 7822 During A Diurnal Light-Dark Cycle, Louis Sherman

Louis A Sherman

Background: Cyanothece sp. PCC 7822 is an excellent cyanobacterial model organism with great potential to be applied as a biocatalyst for the production of high value compounds. Like other unicellular diazotrophic cyanobacterial species, it has a tightly regulated metabolism synchronized to the light-dark cycle. Utilizing transcriptomic and proteomic methods, we were able to quantify the relationships between transcription and translation underlying central and secondary metabolism in response to nitrogen free, 12 hour light and 12 hour dark conditions.

Results: By combining mass-spectrometry based proteomics and RNA-sequencing transcriptomics, we quantitatively measured a total of 6766 mRNAs and 1322 proteins at four …

A Diverse Assemblage Of Indole-3-Acetic Acid Producing Bacteria Associate With Unicellular Green Algae, Christopher Bagwell, Magdalena Piskorska, Tanya Soule, Angela Petelos, Christopher Yeager

Tanya Soule

Microalgae have tremendous potential as a renewable feedstock for the production of liquid transportation fuels. In natural waters, the importance of physical associations and biochemical interactions between microalgae and bacteria is generally well appreciated, but the significance of these interactions to algal biofuels production have not been investigated. Here, we provide a preliminary report on the frequency of co-occurrence between indole-3-acetic acid (IAA)-producing bacteria and green algae in natural and engineered ecosystems. Growth experiments with unicellular algae, Chlorella and Scenedesmus, revealed IAA concentration-dependent responses in chlorophyll content and dry weight. Importantly, discrete concentrations of IAA resulted in cell culture synchronization, …

Function And X-Ray Crystal Structure Of Escherichia Coli Yfde, Elwood A. Mullins, Kelly L. Sullivan, T. Joseph Kappock

T. Joseph Kappock

Many food plants accumulate oxalate, which humans absorb but do not metabolize, leading to the formation of urinary stones. The commensal bacterium Oxalobacter formigenes consumes oxalate by converting it to oxalyl-CoA, which is decarboxylated by oxalyl-CoA decarboxylase (OXC). OXC and the class III CoA-transferase formyl-CoA:oxalate CoA-transferase (FCOCT) are widespread among bacteria, including many that have no apparent ability to degrade or to resist external oxalate. The EvgA acid response regulator activates transcription of the Escherichia coli yfdXWUVE operon encoding YfdW (FCOCT), YfdU (OXC), and YfdE, a class III CoA-transferase that is ~30% identical to YfdW. YfdW and YfdU are necessary …

Cooperative Effects Of Drug-Resistance Mutations In The Flap Region Of Hiv-1 Protease, Jennifer Foulkes-Murzycki, Christina Rosi, Nese Yilmaz, Robert Shafer, Celia Schiffer

Celia A. Schiffer

Understanding the interdependence of multiple mutations in conferring drug resistance is crucial to the development of novel and robust inhibitors. As HIV-1 protease continues to adapt and evade inhibitors while still maintaining the ability to specifically recognize and efficiently cleave its substrates, the problem of drug resistance has become more complicated. Under the selective pressure of therapy, correlated mutations accumulate throughout the enzyme to compromise inhibitor binding, but characterizing their energetic interdependency is not straightforward. A particular drug resistant variant (L10I/G48V/I54V/V82A) displays extreme entropy-enthalpy compensation relative to wild-type enzyme but a similar variant (L10I/G48V/I54A/V82A) does not. Individual mutations of sites …

Physiological Effects Of Nickel Chloride On The Freshwater Cyanobacterium Synechococcus Sp. Iu 625, Brian Nohomovich, Bao T. Nguyen, Michael Quintanilla, Lee H. Lee, Sean R. Murray, Tin-Chun Chu

Tinchun Chu, Ph.D.

Phenotypic And Genotypic Characterization Of Escherichia Coli Isolated From Untreated Surface Waters, Steven L. Daniel, Kai F. Hung, Kristopher J. Janezic, Blake Ferry, Eric W. Hendricks, Brian A. Janiga, Tiffany Johnson, Samantha Murphy, Morgan E. Roberts, Sarah M. Scott, Alexandra N. Theisen

Steven L. Daniel

A common member of the intestinal microbiota in humans and animals is Escherichia coli. Based on the presence of virulence factors, E. coli can be potentially pathogenic. The focus of this study was to isolate E. coli from untreated surface waters (37 sites) in Illinois and Missouri and determine phenotypic and genotypic diversity among isolates. Water samples positive for fecal coliforms based on the Colisure® test were streaked directly onto Eosin Methylene Blue (EMB) agar (37°C) or transferred to EC broth (44.5°C). EC broth cultures producing gas were then streaked onto EMB agar. Forty-five isolates were identified as E. coli …

The Chitobiose Transporter, Chbc, Is Required For Chitin Utilization In Borrelia Burgdorferi, David Nelson

David R. Nelson

Background: The bacterium Borrelia burgdorferi, the causative agent of Lyme disease, is a limited-genome organism that must obtain many of its biochemical building blocks, including N-acetylglucosamine (GlcNAc), from its tick or vertebrate host. GlcNAc can be imported into the cell as a monomer or dimer (chitobiose), and the annotation for several B. burgdorferi genes suggests that this organism may be able to degrade and utilize chitin, a polymer of GlcNAc. We investigated the ability of B. burgdorferi to utilize chitin in the absence of free GlcNAc, and we attempted to identify genes involved in the process. We also examined the …

Evaluation Of Glyphosate-Tolerant Soybean Cultivars For Resistance To Bacterial Pustule, Lopa Goradia, Glen Hartman, Steven L. Daniel

Steven L. Daniel

Xanthomonas axonopodis pv. glycines causes bacterial pustule of soybean, which is a common disease in many soybean-growing areas of the world and is controlled by a single recessive gene (rxp gene) commonly found in many conventional glyphosate-sensitive soybean cultivars. Since glyphosate-tolerant cultivars are commonly planted today, there has been no information about whether these new cultivars have bacterial pustule resistance. The goal of this study was to screen glyphosate-tolerant soybean cultivars for resistance to X. axonopodis pv. glycines. Three experiments were completed to evaluate resistance. In experiment 1, 525 commercial glyphosate-tolerant cultivars from 2001 were inoculated with X. axonopodis pv. …

The Evolution Of Reduced Microbial Killing, Jan A.C. Vriezen Dr., Michael Valliere, Margaret A. Riley Dr.

Jan A.C. Vriezen Dr.

Bacteria engage in a never-ending arms race in which they compete for limited resources and niche space. The outcome of this intense interaction is the evolution of a powerful arsenal of biological weapons. Perhaps the most studied of these are colicins, plasmid-based toxins produced by and active against Escherichia coli. The present study was designed to explore the molecular responses of a colicin-producing strain during serial transfer evolution. What evolutionary changes occur when colicins are produced with no target present? Can killing ability be maintained in the absence of a target? To address these, and other, questions, colicinogenic strains and …

Old Acetogens, New Light, Steven L. Daniel, Harold L. Drake, Anita S. Gößner

Steven L. Daniel

Acetogens utilize the acetyl-CoA Wood-Ljungdahl pathway as a terminal electron-accepting, energy-conserving, CO2-fixing process. The decades of research to resolve the enzymology of this pathway (1) preceded studies demonstrating that acetogens not only harbor a novel CO2-fixing pathway, but are also ecologically important, and (2) overshadowed the novel microbiological discoveries of acetogens and acetogenesis. The first acetogen to be isolated, Clostridium aceticum, was reported by Klaas Tammo Wieringa in 1936, but was subsequently lost. The second acetogen to be isolated, Clostridium thermoaceticum, was isolated by Francis Ephraim Fontaine and co-workers in 1942. C. thermoaceticum became the most extensively studied acetogen and …

The Role Of Sos Boxes In Enteric Bacteriocin Regulation, Jan A.C. Vriezen Dr., Osnat Gillor Dr., Margaret A. Riley Dr.

Jan A.C. Vriezen Dr.

Bacteriocins are a large and functionally diverse family of toxins found in all major lineages of Bacteria. Colicins, those bacteriocins produced by Escherichia coli, serve as a model system for investigations of bacteriocin structure–function relationships, genetic organization, and their ecological role and evolutionary history. Colicin expression is often dependent on host regulatory pathways (such as the SOS system), is usually confined to times of stress, and results in death of the producing cells. This study investigates the role of the SOS system in mediating this unique form of toxin expression. A comparison of all the sequenced enteric bacteriocin promoters reveals …

Impact Of Carbon Sources On Growth And Oxalate Synthesis By The Phytopathogenic Fungus Sclerotinia Sclerotiorum, Bryan J. Culbertson, Jaymie Krone, Erastus Gatebe, Norbert C. Furumo, Steven L. Daniel

Steven L. Daniel

The impact of various supplemental carbon sources (oxalate, glyoxylate, glycolate, pyruvate, formate, malate, acetate, and succinate) on growth and oxalate formation (i.e., oxalogenesis) by Sclerotinia sclerotiorum was studied. With isolates D-E7, 105, W-B10, and Arg-L of S. sclerotiorum, growth in an undefined broth medium (0.1% soytone; pH 5) with 25 mM glucose and 25 mM supplemental carbon source was increased by the addition of malate and succinate. Oxalate accumulation occurred in the presence of glucose and a supplemental carbon source, with malate, acetate, and succinate supporting the most oxalate synthesis. With S. sclerotiorum Arg-L, oxalate-to-biomass ratios, an indicator of oxalogenic …

Mammalian Cell Cytotoxicity Analysis Of Soybean Rust Fungicides, Steven L. Daniel, G. L. Hartman, E. D. Wagner, M. J. Plewa

Steven L. Daniel

No abstract provided.

Impact Of Nutritional Supplements And Monosaccharides On Growth, Oxalate Accumulation, And Culture Ph By Sclerotinia Sclerotiorum, Steven L. Daniel, Bryan J. Culbertson, Norbert C. Furumo

Steven L. Daniel

Sclerotinia sclerotiorum D-E7 was studied to determine the impact of nutritional supplements and monosaccharides on growth, oxalate accumulation, and culture pH in broth media (initial pH c. 5). Cultures with 0.1% nutritional supplement (tryptone, yeast extract, or soytone) yielded minimal growth, 2–3 mM oxalate, and a final culture pH of 4.2–4.8. In contrast, cultures with 0.1% nutritional supplement and 25 mM glucose yielded significant growth, minimal oxalate (<1 mM), and a final culture pH of 2.8–3.7. Similar trends were observed when glucose in 0.1% soytone cultures was replaced with 25 mM d-mannose, l-arabinose, or d-xylose. With 1% soytone-25 mM glucose cultures, growth and oxalate accumulation (∼21 mM) occurred with little change in initial pH. This was not the case with 1% soytone-250 mM glucose cultures; increased glucose levels resulted in a decrease in oxalate accumulation (∼7 mM) and in final culture pH (3.4). Time-course studies with these cultures revealed that oxalate accumulation was suppressed during growth when the culture pH dropped to <4. Overall, these results indicate that (1) the decrease in external pH (i.e. acidification) was independent of oxalate accumulation and (2) acidification coupled to glucose-dependent growth regulated oxalate accumulation by Sclerotinia sclerotiorum.

Anaerobic Oxalate Consumption By Microorganisms In Forest Soils, Steven L. Daniel, Christine Pilsl, Harold L. Drake

Steven L. Daniel

The microbial consumption of oxalate was examined under anaerobic conditions in soil suspensions at 15-20 degree C. With soil (horizon Ah, pH 6.4) from a beech forest, microbial consumption of added oxalate (15 mM) began after 10 days, and oxalate was totally consumed by day 20. The presence of supplemental electron donors (acetate, glucose, vanillate, or hydrogen) or electron acceptors (nitrate or sulfate) did not significantly influence anaerobic oxalate consumption, whereas supplementation of soil suspensions with CO2/bicarbonate totally repressed oxalate consumption. Thus, CO2-, nitrate- or sulfate-respiring bacteria were apparently not active in the anaerobic consumption of oxalate in these soil …

Physiology Of The Thermophilic Acetogen Moorella Thermoacetica, Harold L. Drake, Steven L. Daniel

Steven L. Daniel

Moorella thermoacetica (originally isolated as Clostridium thermoaceticum) has served as the primary acetogenic bacterium for the resolution of the acetyl coenzyme A (acetyl-CoA) orWood–Ljungdahl pathway, a metabolic pathway that (i) autotrophically assimilates CO2 and (ii) is centrally important to the turnover of carbon in many habitats. The purpose of this article is to highlight the diverse physiological features of this model acetogen and to examine some of the consequences of its metabolic capabilities.

Oxalate Metabolism By The Acetogenic Bacterium Moorella Thermoacetica, Steven L. Daniel, Christine Pilsl, Harold L. Drake

Steven L. Daniel

Whole-cell and cell-extract experiments were performed to study the mechanism of oxalate metabolism in the acetogenic bacterium Moorella thermoacetica. In short-term, whole-cell assays, oxalate consumption was low unless cell suspensions were supplemented with CO2, KNO3, or Na2S2O3. Cell extracts catalyzed the oxalate-dependent reduction of benzyl viologen. Oxalate consumption occurred concomitant to benzyl viologen reduction; when benzyl viologen was omitted, oxalate was not appreciably consumed. Based on benzyl viologen reduction, specific activities of extracts averaged 0.6 μmol oxalate oxidized min−1 mg protein−1. Extracts also catalyzed the formate-dependent reduction of NADP+; however, oxalate-dependent reduction of NADP+ was negligible. Oxalate- or formate-dependent reduction …

A New Genetic Locus In Sinorhizobium Meliloti Is Involved In Stachydrine Utilization, Donald A. Phillips Dr., Eve S. Sandee Dr., Jan A.C. Vriezen Dr., Frans J. Debruijn Dr., Daniel Lerudulier Dr., Cecillia M. Joseph Dr.

Jan A.C. Vriezen Dr.

Stachydrine, a betaine released by germinating alfalfa seeds, functions as an inducer of nodulation genes, a catabolite, and an osmoprotectant in Sinorhizobium meliloti. Two stachydrine-inducible genes were found in S. meliloti 1021 by mutation with a Tn5-luxAB promoter probe. Both mutant strains (S10 and S11) formed effec- tive alfalfa root nodules, but neither grew on stachydrine as the sole carbon and nitrogen source. When grown in the absence or presence of salt stress, S10 and S11 took up [14C]stachydrine as well as wild-type cells did, but neither used stachydrine effectively as an osmoprotectant. In the absence of salt stress, both …

Microbial Degradation Of Oxalate In The Gastrointestinal Tracts Of Rats, Steven L. Daniel

Steven L. Daniel

Rates of oxalate degradation by mixed bacterial populations in cecal contents from wlld rats ranged from 2.5 to 20.6 µmol/g (dry weight) per h. The oxalate-degrading activity in cecal contents from three strains of laboratory rats (Long-Evans, Wistar, and Sprague-Dawley) from four commercial breeders was generally lower, ranging from 1.8 to 3.5 µmollg (dry weight) of cecal contents per h. This activity did not increase when diets were supplemented with oxalate. Wben Sprague-Pawley rats from a fifth commercial breeder were fed an oxalate diet, rates of oxalate degradation in cecal contents increased from 2.0 to 23.1 µmollg (dry weight) per …

Carbon Monoxide-Dependent Chemolithotrophic Growth Of Clostridium Thermoautotrophicum, M. Dean Savage, Zhongren Wu, Steven L. Daniel, Leon L. Lundie Jr, Harold L. Drake

Steven L. Daniel

The acetogen Clostridium thermoautotrophicum was cultivated under CO-dependent chemolithotrophic conditions. CO-dependent growth profiles and energetics indicated that supplemental CO2 was fundamental to efficient growth at the expense of CO. Overall product stoichiometry approximated 6.5CO --> CH3CO2H + 3.5CO2 + 0.6 cell C + 0.5 unrecovered C. Initial CO/CO2 ratios of 2 to 4 yielded optimal doubling times and cell yields. Maximal YCO values approximated 2.5 g of cell dry weight per mol of CO consumed; Y H2 , was considerably lower than Y CO Cross-transfer growth experiments and protein profiles indicated differential expression of genes between CO and methanol cultures.