Microbiology Commons^™

Roles Of Three Transporters, Cbcxwv, Bett1, And Bett3, In Pseudomonas Aeruginosa Choline Uptake For Catabolism, Adel A. Malek, Chiliang Chen, Matthew J. Wargo, Gwyn A. Beattie, Deborah A. Hogan

Dartmouth Scholarship

Pseudomonas aeruginosa uses the quaternary amine choline as a carbon source, osmoprotectant, and macromolecular precursor. The importance of choline in P. aeruginosa physiology is highlighted by the presence of multiple known and putative choline transporters encoded within its genome. This report describes the relative roles of three choline transporters, the ABC transporter CbcXWV and two symporters, BetT1 and BetT3, in P. aeruginosa growth on choline under osmotic conditions that are physiologically relevant to eukaryotic hosts. The increased lag phases exhibited by the ΔbetT1 and ΔbetT1 ΔbetT3 mutants relative to the wild type upon transfer to medium with …

Whole-Genome Sequencing Of Staphylococcus Aureus Strain Rn4220, A Key Laboratory Strain Used In Virulence Research, Identifies Mutations That Affect Not Only Virulence Factors But Also The Fitness Of The Strain, Dhanalakshmi Nair, Guido Memmi, David Hernandez, Jonathan Bard, Marie Beaume, Steven Gill, Patrice Francois, Ambrose L. Cheung

Dartmouth Scholarship

Staphylococcus aureus RN4220, a cloning intermediate, is sometimes used in virulence, resistance, and metabolic studies. Using whole-genome sequencing, we showed that RN4220 differs from NCTC8325 and contains a number of genetic polymorphisms that affect both virulence and general fitness, implying a need for caution in using this strain for such studies.

Pcr Primers For The Detection Of Propane And Butane-Oxidizing Microorganisms, Brian Jeremy Chan

Master's Theses

In an increasingly energy-hungry world, our capacity to meet the heightened energy demands of the future has become a pressing matter. The most urgent of these concerns are tied to the accessibility of petroleum. Various experts have proselytized both the imminent arrival of peak oil production rates and the ensuing decline of those rates thereafter. And to that end, the development of novel and advanced oil exploration methodologies has become almost as important as finding the sources of oil themselves.

The soils above petroleum reservoirs play host to various communities of alkane- oxidizing bacteria that can utilize the natural gas …

Cellulose- And Xylan-Degrading Thermophilic Anaerobic Bacteria From Biocompost, M. V. Sizova, J. A. Izquierdo, N. S. Panikov, L. R. Lynd

Dartmouth Scholarship

Nine thermophilic cellulolytic clostridial isolates and four other noncellulolytic bacterial isolates were isolated from self-heated biocompost via preliminary enrichment culture on microcrystalline cellulose. All cellulolytic isolates grew vigorously on cellulose, with the formation of either ethanol and acetate or acetate and formate as principal fermentation products as well as lactate and glycerol as minor products. In addition, two out of nine cellulolytic strains were able to utilize xylan and pretreated wood with roughly the same efficiency as for cellulose. The major products of xylan fermentation were acetate and formate, with minor contributions of lactate and ethanol. Phylogenetic analyses of 16S …

Laser-Induced Breakdown Spectroscopy (Libs): An Innovative Tool For Studying Bacteria, Qassem I. Mohaidat

Wayne State University Dissertations

Laser-induced breakdown spectroscopy (LIBS) has gained a reputation as a flexible and convenient technique for rapidly determining the elemental composition of samples with minimal or no sample preparation. In this dissertation, I will describe the benefits of using LIBS for the rapid discrimination and identification of bacteria (both pathogenic and non-pathogenic) based on the relative concentration of trace inorganic elements such as Mg, P, Ca, and Na. The speed, portability, and robustness of the technique suggest that LIBS may be applicable as a rapid point-of-care medical diagnostic technology.

LIBS spectra of multiple genera of bacteria such as Escherichia, Streptococcus, Mycobacterium, …

Microbial Landscapes Of Corals And Ctenophores, Camille Arian Daniels

USF Tampa Graduate Theses and Dissertations

As technology and engineering allow mankind to survey nature at finer scales, the importance of bacteria has been elucidated in their metabolic diversity, ability to transfer genetic information, involvement in biogeochemical cycling, and sheer abundance. With an individual domain of life unto themselves, this diverse group of microorganisms plays an integral role in facilitating life on land and in the oceans, and is second only to viruses in abundance on Earth. They carve niches in a wide range of environments, including those inhospitable to other life forms, and reside in concert or to the detriment of other microbes and/or hosts …

Development Of Pyrf-Based Genetic System For Targeted Gene Deletion In Clostridium Thermocellum And Creation Of A Pta Mutant, Shital A. Tripathi, Daniel G. Olson, D. Aaron Argyros, Bethany B. Miller, Trisha F. Barrett, Daniel M. Murphy, Jesse D. Mccool, Anne K. Warner, Vineet B. Rajgarhia, Lee R. Lynd, David A. Hogsett, Nicky C. Caiazza

Dartmouth Scholarship

We report development of a genetic system for making targeted gene knockouts in Clostridium thermocellum, a thermophilic anaerobic bacterium that rapidly solubilizes cellulose. A toxic uracil analog, 5-fluoroorotic acid (5-FOA), was used to select for deletion of the pyrF gene. The ΔpyrF strain is a uracil auxotroph that could be restored to a prototroph via ectopic expression of pyrF from a plasmid, providing a positive genetic selection. Furthermore, 5-FOA was used to select against plasmid-expressed pyrF, creating a negative selection for plasmid loss. This technology was used to delete a gene involved in organic acid production, namely pta, which encodes …

Isolation And Characterization Of Bacteria From Extended Aeration Wastewater Treatment Plants., Randy Chin Kok Fei

Student Works (2010-2019)

In this study, three different wastewater treatment plants (STP1, STP2 and STP3) were sampled in Klang Valley. STP1 exhibited the best activated sludge health with healthy Sludge Volume (SV), Mixed Liquor Suspended Solids (MLSS) and Mixed Liquor Volatile Suspended Solids (MLVSS). STP2 were constantly disrupted by the mechanical failures which caused its dissolved oxygen (DO) to decrease whereas STP3 was troubled by the inadequate alkalinity in its mixed liquor causing acidic condition in aeration tank. The acidic condition resulted in pin-floc problem and contributed to high Total Suspended Solids (TSS) and low SV. Influent at STP3 was also found to …

Copper Isotope Fractionation During Surface Adsorption And Intracellular Incorporation By Bacteria, Jesica Urbina Navarrete

Open Access Theses & Dissertations

Copper isotopes may prove to be a useful tool for investigating bacteria-metal interactions recorded in natural waters, soils, and rocks. However, experimental data that constrain Cu isotope fractionation in biologic systems are limited and unclear. In this study we utilized Cu isotopes (65Cu) as a tool to investigate Cu-bacteria interactions, including surface adsorption and intracellular incorporation. Experiments were conducted with individual Gram-positive (Bacillus subtilis) and Gram-negative (Escherichia coli) bacterial species as well as with bacterial consortia from several natural environments. Adsorption experiments were conducted with live or dead cells over the pH range 2.5 to 6. Surface adsorption of Cu …

Proteolytic Regulation Of Toxin-Antitoxin Systems By Clppc In Staphylococcus Aureus, Niles P. Donegan, Earl T. Thompson, Zhibiao Fu, Ambrose L. Cheung

Dartmouth Scholarship

Bacterial toxin-antitoxin (TA) systems typically consist of a small, labile antitoxin that inactivates a specific longer-lived toxin. In Escherichia coli, such antitoxins are proteolytically regulated by the ATP-dependent proteases Lon and ClpP. Under normal conditions, antitoxin synthesis is sufficient to replace this loss from proteolysis, and the bacterium remains protected from the toxin. However, if TA production is interrupted, antitoxin levels decrease, and the cognate toxin is free to inhibit the specific cellular component, such as mRNA, DnaB, or gyrase. To date, antitoxin degradation has been studied only in E. coli, so it remains unclear whether similar mechanisms of regulation …

Levels Of The Secreted Vibrio Cholerae Attachment Factor Gbpa Are Modulated By Quorum-Sensing-Induced Proteolysis, Brooke A. Jude, Raquel M. Martinez, Karen Skorupski, Ronald K. Taylor

Dartmouth Scholarship

Vibrio cholerae is the etiologic agent of cholera in humans. Intestinal colonization occurs in a stepwise fashion, initiating with attachment to the small intestinal epithelium. This attachment is followed by expression of the toxin-coregulated pilus, microcolony formation, and cholera toxin (CT) production. We have recently characterized a secreted attachment factor, GlcNAc binding protein A (GbpA), which functions in attachment to environmental chitin sources as well as to intestinal substrates. Studies have been initiated to define the regulatory network involved in GbpA induction. At low cell density, GbpA was detected in the culture supernatant of all wild-type (WT) strains examined. In …

Identification Of The [Fefe]-Hydrogenase Responsible For Hydrogen Generation In Thermoanaerobacterium Saccharolyticum And Demonstration Of Increased Ethanol Yield Via Hydrogenase Knockout, A. Joe Shaw, David A. Hogsett, Lee R. Lynd

Dartmouth Scholarship

Three putative hydrogenase enzyme systems in Thermoanaerobacterium saccharolyticum were investigated at the genetic, mRNA, enzymatic, and phenotypic levels. A four-gene operon containing two [FeFe]-hydrogenase genes, provisionally termed hfs (hydrogenase-Fe-S), was found to be the main enzymatic catalyst of hydrogen production. hfsB, perhaps the most interesting gene of the operon, contains an [FeFe]-hydrogenase and a PAS sensory domain and has several conserved homologues among clostridial saccharolytic, cellulolytic, and pathogenic bacteria. A second hydrogenase gene cluster, hyd, exhibited methyl viologen-linked hydrogenase enzymatic activity, but hyd gene knockouts did not influence the hydrogen yield of …

Microbial Nad Metabolism: Lessons From Comparative Genomics, Francesca Gazzaniga, Rebecca Stebbins, Sheila Z. Chang, Mark A. Mcpeek, Charles Brenner

Dartmouth Scholarship

NAD is a coenzyme for redox reactions and a substrate of NAD-consuming enzymes, including ADP-ribose transferases, Sir2-related protein lysine deacetylases, and bacterial DNA ligases. Microorganisms that synthesize NAD from as few as one to as many as five of the six identified biosynthetic precursors have been identified. De novo NAD synthesis from aspartate or tryptophan is neither universal nor strictly aerobic. Salvage NAD synthesis from nicotinamide, nicotinic acid, nicotinamide riboside, and nicotinic acid riboside occurs via modules of different genes. Nicotinamide salvage genes nadV and pncA, found in distinct bacteria, appear to have spread throughout the tree of life …

Pseudomonas Aeruginosa-Candida Albicans Interactions: Localization And Fungal Toxicity Of A Phenazine Derivative, Jane Gibson, Arpanah Sood, Deborah A. Hogan

Dartmouth Scholarship

Phenazines are redox-active small molecules that play significant roles in the interactions between pseudomonads and diverse eukaryotes, including fungi. When Pseudomonas aeruginosa and Candida albicans were cocultured on solid medium, a red pigmentation developed that was dependent on P. aeruginosa phenazine biosynthetic genes. Through a genetic screen in combination with biochemical experiments, it was found that a P. aeruginosa-produced precursor to pyocyanin, proposed to be 5-methyl-phenazinium-1-carboxylate (5MPCA), was necessary for the formation of the red pigmentation. The 5MPCA-derived pigment was found to accumulate exclusively within fungal cells, where it retained the ability to be reversibly oxidized and reduced, and its …

The Effects Of Host Physiological Conditions On The Expression Of Icsp In Shigella Flexneri, Karen Levy, Helen Wing

Undergraduate Research Opportunities Program (UROP)

Shigella flexneri is a gram-negative bacterium capable of causing diarrhea and dysentery known as shigellosis. It is estimated there are 167.4 million shigellosis episodes throughout the world each year causing 1.1 million deaths. Shigella invades cells in the lower intestine through an induced phagocytosis. Once in the cytoplasm, bacteria move from one cell to another using actin-based motility. The Shigella outer membrane protease IcsP regulates actin-based motility and cell-to-cell spread by cleaving the actin assembly protein IcsA from the bacterial cell surface. We hypothesize that IcsP may serve additional functions during infection. By examining which environmental signals trigger icsP expression, …

Microbial Ecology Of Keane Wonder Spring, Death Valley National Park, Alexander B. Michaud, Duane P. Moser

Undergraduate Research Opportunities Program (UROP)

This research is focused on developing a better understanding of he physiological and phylogenetic diversity as well as environmental abundance of bacteria of the genus: Shewanella in selected desert ecosystems. Prior research from this laboratory has revealed that these bacteria are very abundant in sulfurand organic-rich aquatics habitats. We have selected a number of habitats for detailed investigation (cultivation, molecular ecology and relevant environmental chemistry) including the Tropicana Wash, spring in Death Valley, the lower Virgin River and possibly Big Soda Lame, Nevada.

The Regulation Of The Icsp Promoter Of Shigella Flexneri By The Virulence Factor Virb, Maria Castellanos, Dustin Harrison, Helen Wing

Undergraduate Research Opportunities Program (UROP)

Shigella flexneri is a pathogenic bacterium that causes severe dysentery in humans commonly known as shigellosis. Shigella encodes an outer membrane protease called IcsP. The regulation of icsP expression is under direct control of a transcriptional factor called VirB, which controls the expression of many virulence genes in Shigella. Previous work has shown through deletion analysis of the icsP promoter region that sequences as far as 1368 base pairs upstream of the transcription starting site are important for the regulation of the icsP gene by VirB. However, it is still unclear whether VirB activation requires sequences within the icsP promoter …

Characterization Of The Omptin Protease, Ompt, In Escherichia Coli, Amanda Yates, Eun-Hae Kim, Helen Wing

Undergraduate Research Opportunities Program (UROP)

Omptins are outer membrane proteases found in gram negative bacteria that cause diseases in humans, such as pathogenic Escherichia coli, Shigella flexneri, Salmonella typhimurium, and Yersinia pestis. Bacterial species that express omptins cause diseases such as highly fatal plague and severe diarrhea and dysentery. The genes that encode these proteases are ompT, icsP, pgtE, and pla, respectively. These proteases are highly related in structure and share approximately 50% sequence identity. In S. flexneri, IcsP has been shown to cleave a key virulence determinant, IcsA (Egile et al., 1997). IcsA recruits host actin and allows for intracellular movement within host cells …

Sara Of Staphylococcus Aureus Binds To The Sara Promoter To Regulate Gene Expression, Ambrose L. Cheung, Koren Nishina, Adhar C. Manna

Dartmouth Scholarship

The 375-bp sarA open reading frame is driven by three promoters, P1, P3, and P2. Using gel shift and DNase I footprinting assays, we found that SarA binds to two 26-bp sequences and one 31-bp sequence within the P1 and P3 promoters, respectively. Together with the results of transcription analyses, our data indicate that SarA binds to its own promoter to down-regulate sarA expression.

Antimicrobial Activity Screening Of Recombinant And Synthetic Varasin A Defensin From The Hard Tick Dermacentor Variabilis Against Various Bacteria, Julia A. Sharp

Biological Sciences Theses & Dissertations

Hematophagous arthropods, such as ticks and mosquitoes, rely on their innate immune system for defense against pathogens ingested in a blood meal as well as those acquired through injury. In response to pathogen recognition, the production of antimicrobial peptides, such as defensin, is typically upregulated. Varisin, a defensin, is thought to be a key component in the immunocompetence of the hard tick D. variabilis against Borrelia burgdorferi, the causative agent of Lyme disease. To study the antimicrobial effects of varisin, recombinant varisin was expressed by both insect cells and E coli. Purification of the protein followed by enterokinase treatment yielded …

Environmental Controls On The Landscape-Scale Biogeography Of Stream Bacterial Communities, Noah Fierer, Jennifer L. Morse, Sean T. Berthrong, Emily S. Bernhardt, Robert B. Jackson

Environmental Science and Management Faculty Publications and Presentations

We determined the biogeographical distributions of stream bacteria and the biogeochemical factors that best explained heterogeneity for 23 locations within the Hubbard Brook watershed, a 3000-ha forested watershed in New Hampshire, USA. Our goal was to assess the factor, or set of factors, responsible for generating the biogeographical patterns exhibited by microorganisms at the landscape scale. We used DNA fingerprinting to characterize bacteria inhabiting fine benthic organic matter (FBOM) because of their important influence on stream nutrient dynamics. Across the watershed, streams of similar pH had similar FBOM bacterial communities. Streamwater pH was the single variable most strongly correlated with …

Characterization Of Dairy Milk House Waste Water In Kentucky, Anshu Singh, Czarena L. Crofcheck, Gail M. Brion

Biosystems and Agricultural Engineering Faculty Publications

This study focuses on characterization of milk house waste water from eight different farms in Kentucky. The farms were separated into three groups based on the number of cows: small (20-30), medium (30-60), and large (over 60 cows). Samples were collected once a month from four farms and twice a month from the remainder. Samples were analyzed for chemical, biochemical, and microbiological characteristics. Results indicated a large and significant variation in the chemical and microbiological characteristics between the farms. Farm size had a significant effect on the nutrient content of the waste water. Though samples exhibited seasonal variation, there was …

Environmental Controls On The Landscape-Scale Biogeography Of Stream Bacterial Communities, Noah Fierer, Jennifer L. Morse, Sean T. Berthrong, Emily S. Bernhardt, Robert B. Jackson

Scholarship and Professional Work - LAS

We determined the biogeographical distributions of stream bacteria and the biogeochemical factors that best explained heterogeneity for 23 locations within the Hubbard Brook watershed, a 3000-ha forested watershed in New Hampshire, USA. Our goal was to assess the factor, or set of factors, responsible for generating the biogeographical patterns exhibited by microorganisms at the landscape scale. We used DNA fingerprinting to characterize bacteria inhabiting fine benthic organic matter (FBOM) because of their important influence on stream nutrient dynamics. Across the watershed, streams of similar pH had similar FBOM bacterial communities. Streamwater pH was the single variable most strongly correlated with …

Good Things Come In Small Packages: Tiny Plankton Producing Oxygen Near Dead Zone, Abbie Basile

Libraries Faculty & Staff Publications

(First paragraph) For many, the words “Lake Erie” bring to mind large things: big sport fish, sprawling waterfront homes, and an enormous body of water bordering four states and two countries. Similarly, much of the Lake Erie research of which the public is aware deals with larger biological organisms—larger algae and zooplankton, the small fish that eat the zooplankton, and the large fish that feed on those smaller fish. Over the years, the smaller lake life have been somewhat overlooked.

Conservation Of The Pho Regulon In Pseudomonas Fluorescens Pf0-1, Russell D. Monds, Peter D. Newell, Julia A. Schwartzman, George A. O'Toole

Dartmouth Scholarship

The Pho regulon integrates the sensing of environmental inorganic phosphate (Pi) availability with coregulation of gene expression, mediating an adaptive response to Pi limitation. Many aspects of the Pho regulon have been addressed in studies of Escherichia coli; however, it is unclear how transferable this knowledge is to other bacterial systems. Here, we report work to discern the conservation of the Pho regulon in Pseudomonas fluorescens Pf0-1. We demonstrate by mutational studies that PhoB/PhoR and the Pst system have conserved functions in the regulation of Pi-induced phosphatase activities, as well as expression of other Pi-regulated genes. A genetic screen was …

Transposon Disruption Of The Complex I Nadh Oxidoreductase Gene (Snod) In Staphylococcus Aureus Is Associated With Reduced Susceptibility To The Microbicidal Activity Of Thrombin-Induced Platelet Microbicidal Protein 1, Arnold S. Bayer, Peter Mcnamara, Michael R. Yeaman, Natalie Lucindo, Tiffanny Jones, Ambrose L. Cheung

Dartmouth Scholarship

The cationic molecule thrombin-induced platelet microbicidal protein 1 (tPMP-1) exerts potent activity against Staphylococcus aureus. We previously reported that a Tn551 S. aureus transposon mutant, ISP479R, and two bacteriophage back-transductants, TxA and TxB, exhibit reduced in vitro susceptibility to tPMP-1 (tPMP-1(r)) compared to the parental strain, ISP479C (V. Dhawan, M. R. Yeaman, A. L. Cheung, E. Kim, P. M. Sullam, and A. S. Bayer, Infect. Immun. 65:3293-3299, 1997). In the current study, the genetic basis for tPMP-1(r) in these mutants was identified. GenBank homology searches using sequence corresponding to chromosomal DNA flanking Tn551 mutant strains showed that the fourth gene …

Rhamnolipids Modulate Swarming Motility Patterns Of Pseudomonas Aeruginosa, Nicky C. Caiazza, Robert M. Q. Shanks, G. A. O'Toole

Dartmouth Scholarship

Pseudomonas aeruginosa is capable of twitching, swimming, and swarming motility. The latter form of translocation occurs on semisolid surfaces, requires functional flagella and biosurfactant production, and results in complex motility patterns. From the point of inoculation, bacteria migrate as defined groups, referred to as tendrils, moving in a coordinated manner capable of sensing and responding to other groups of cells. We were able to show that P. aeruginosa produces extracellular factors capable of modulating tendril movement, and genetic analysis revealed that modulation of these movements was dependent on rhamnolipid biosynthesis. An rhlB mutant (deficient in mono- and dirhamnolipid production) and …

Role Of Spontaneous Current Oscillations During High-Efficiency Electrotransformation Of Thermophilic Anaerobes, Michael V. Tyurin, Charles R. Sullivan, Lee R. Lynd

Dartmouth Scholarship

Current oscillations at about 24 MHz were observed during electrotransformation (ET) of the thermophilic anaerobes Clostridium thermocellum ATCC 27405, C. thermocellum DSM 1313, and Thermoanaerobacterium saccharolyticum YS 485, using a pulse gated by a square signal generated by a custom generator. In experiments in which only the field strength was varied, all three of these strains resulted in a one-to-one correspondence between the appearance of current oscillations and successful ET. Oscillations accompanied ET of both C. thermocellum strains only at field strengths of >12 kV/cm, and ET was only observed above the same threshold. Similarly, for T. saccharolyticum, oscillations were …

Requirements For Vibrio Cholerae Hapr Binding And Transcriptional Repression At The Hapr Promoter Are Distinct From Those At The Apha Promoter, Wei Lin, Gabriela Kovacikova, Karen Skorupski

Dartmouth Scholarship

Virulence gene expression in certain strains of Vibrio cholerae is regulated in response to cell density by a quorum-sensing cascade that influences the levels of the LuxR homolog HapR through small regulatory RNAs that control the stability of its message. At high cell density, HapR represses the expression of the gene encoding the virulence gene activator AphA by binding to a site between −85 and −58 in the aphA promoter. We show here that a second binding site for HapR lies within the hapR promoter from which it functions to repress its own transcription. This site, as determined by gel …

Identification Of A Tcpc-Tcpq Outer Membrane Complex Involved In The Biogenesis Of The Toxin-Coregulated Pilus Of Vibrio Cholerae, Niranjan Bose, Ronald K. Taylor

Dartmouth Scholarship

The toxin-coregulated pilus (TCP) of Vibrio cholerae and the soluble TcpF protein that is secreted via the TCP biogenesis apparatus are essential for intestinal colonization. The TCP biogenesis apparatus is composed of at least nine proteins but is largely uncharacterized. TcpC is an outer membrane lipoprotein required for TCP biogenesis that is a member of the secretin protein superfamily. In the present study, analysis of TcpC in a series of strains deficient in each of the TCP biogenesis proteins revealed that TcpC was absent specifically in a tcpQ mutant. TcpQ is a predicted periplasmic protein required for TCP biogenesis. Fractionation …