Microbiology Commons^™

Cloning And Functional Analysis Of The Escherichia Coli Cell Division Protein Zape, Katherine Kellenberger

Senior Honors Projects

During bacterial cell division, a large, dynamic cytoskeletal structure called the Z-ring assembles at the site of division. The Z-ring is comprised of the major cell division protein FtsZ, a tubulin-like GTPase that utilizes GTP to assemble into linear polymers. In Escherichia coli, there are several cell division proteins that interact with FtsZ and regulate Z-ring assembly, constriction, and disassembly. The accessory proteins that interact with FtsZ are called Z-ring associated proteins (ZAPs). The Zaps which include ZapA, ZapB, ZapC, ZapD and ZapE are recruited to the divisome and influence Z-ring assembly and stability. Specifically, ZapE was identified to be …

The Role Of Genomic Versatility In Multi-Niche Preferences Of Escherichia Coli, Gitanjali Nandakafle

Electronic Theses and Dissertations

Escherichia coli strains are naturally present as either commensals or pathogens in the gastrointestinal tract of mammals and some other vertebrates. Until recently, it was assumed that E. coli are solely associated with the gut and are unable to survive outside of a host for a long period of time, the basis of its use as an indicator organism. Recent reports suggest that E. coli can become naturalized to several tropical, subtropical or temperate soils and aquatic environments, where they have been isolated repeatedly. Several studies have shown that these strains are capable of surviving and proliferating in the environment …

Are Cdi Systems Multicolored, Facultative, Helping Greenbeards?, Elizabeth S. Danka, Erin C. Garcia, Peggy A. Cotter

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Competitive and cooperative interactions between organisms, including bacteria, can significantly impact the composition of a community and the fitness of its members, as well as the fitness of their hosts when communities are living on or within other organisms. Understanding the underlying mechanisms is critical to the development of strategies to control microbiological communities that impact animal and plant health and also for understanding the evolution of social behaviors, which has been challenging for evolutionary biologists. Contact-dependent growth inhibition (CDI) is a phenomenon defined by the delivery of a protein toxin to the cytoplasm of neighboring bacteria upon cell–cell contact, …

Genomic Analysis Of Factors Associated With Low Prevalence Of Antibiotic Resistance In Extraintestinal Pathogenic Escherichia Coli Sequence Type 95 Strains, Craig M. Stephens, Sheila Adams-Sapper, Manraj Sekhon, James R. Johnson, Lee W. Riley

Biology

Extraintestinal pathogenic Escherichia coli (ExPEC) strains belonging to multilocus sequence type 95 (ST95) are globally distributed and a common cause of infections in humans and domestic fowl. ST95 isolates generally show a lower prevalence of acquired antimicrobial resistance than other pandemic ExPEC lineages. We took a genomic approach to identify factors that may underlie reduced resistance. We fully assembled genomes for four ST95 isolates representing the four major fimH-based lineages within ST95 and also analyzed draft-level genomes from another 82 ST95 isolates, largely from the western United States. The fully assembled genomes of antibiotic-resistant isolates carried resistance genes exclusively on …

Mutagenic And Spectroscopic Investigation Of Ph Dependent Cooa Dna Binding, Brian R. Weaver

Chemistry Honors Papers

The carbon monoxide (CO) sensing heme protein, CooA, is a transcription factor which exists in several bacteria that utilize CO as an energy source. CooA positively regulates the expression of coo genes in the presence of CO such that the corresponding proteins may metabolize CO. The present studies have yielded the unexpected result that Fe(III) CooA binds DNA tightly at pH < 7, deviating from all previously reported work which indicate that CooA DNA binding is initiated only when the exogenous CO effector reacts with the Fe(II) CooA heme. This observation suggests that the disruption of one or more salt bridges upon effector binding may be a critical feature of the normal CooA activation mechanism. To test this possibility, several protein variants that eliminated a selected salt bridge for the CooA homolog from Rhodospirillum rubrum were prepared via site-directed mutagenesis. Samples of these variant proteins, which were overexpressed in Escherichia coli, were then characterized by spectroscopic methods and functional assays to investigate the impact these mutations had on CooA heme coordination …

Computing And Applying Atomic Regulons To Understand Gene Expression And Regulation, Jose P. Faria, James J. Davis, Janaka N. Edirisinghe, Ronald C. Taylor, Pamela B. Weisenhorn, Robert D. Olson, Rick Stevens, Miguel Rocha, Isabel Rocha, Aaron A. Best, Matthew Dejongh, Nathan L. Tintle, Bruce Parrelo, Ross Overbeek, Christopher S. Henry

Faculty Work Comprehensive List

Understanding gene function and regulation is essential for the interpretation prediction and ultimate design of cell responses to changes in the environment. An important step toward meeting the challenge of understanding gene function and regulation is the identification of sets of genes that are always co-expressed. These gene sets Atomic Regulons ARs represent fundamental units of function within a cell and could be used to associate genes of unknown function with cellular processes and to enable rational genetic engineering of cellular systems. Here we describe an approach for inferring ARs that leverages large-scale expression data sets gene context and functional …

Genome Sequences And Annotation Of Two Urinary Isolates Of E.Coli, Travis Kyle Price, Arya Mehtash, Laurynas Kalesinskas, Kema Malki, Evann Elizabeth Hilt, Catherine Putonti, Alan J. Wolfe

Bioinformatics Faculty Publications

The genus Escherichia includes pathogens and commensals. Bladder infections (cystitis) result most often from colonization of the bladder by uropathogenic E. coli strains. In contrast, a poorly defined condition called asymptomatic bacteriuria results from colonization of the bladder with E. coli strains without symptoms. As part of an on-going attempt to identify and characterize the newly discovered female urinary microbiota, we report the genome sequences and annotation of two urinary isolates of E. coli: one (E78) was isolated from a female patient who self-reported cystitis; the other (E75) was isolated from a female patient who reported that she did …

Contribution Of A Putative Up Element Dna Sequence To The Activity Of A Newly Identified Phage Promoter, Courtney Hamilton

Mahurin Honors College Capstone Experience/Thesis Projects

In transcription, a universal step in gene expression, information from a DNA sequence is copied into RNA. A key component in gene expression is the promoter sequence, a region of DNA to which RNA polymerase binds during the initiation of transcription of downstream genes. Most bacterial promoters contain a -10 and a -35 sequence that are bound by the RNA polymerase. Some promoters also contain an Upstream Promoter (UP) element. UP elements have been shown to boost promoter activity. We recently identified a new promoter in a mutant bacteriophage that grows on a bacterial host that prevents antitermination of phage …

Assessing Stress Responses To Atmospheric Cold Plasma Exposure Using Escherichia Coli Knock Out Mutants, Lu Han, Daniela Boehm, Sonal Patil, Patrick J. Cullen, Paula Bourke

Articles

Aim: This study investigated the effect of Atmospheric cold plasma (ACP) exposure induced stress on microbial inactivation patterns and the regulation of genes involved in the microbial stress response in conjunction with key processing parameters of exposure time and post treatment storage time.

Methods and Results: Cell suspensions of Escherichia coli BW 25113 and its isogenic knock-out mutants in rpoS, soxR, soxS, oxyR and dnaK genes were treated with high voltage ACP in a sealed package for 1, 3 and 5 min followed by 0, 1 and 24 h post-treatment storage. ROS densities and colony formation were …

Detection Methods And Intestinal Adherence Of Non-O157 Shiga Toxin-Producing Escherichia Coli, Zachary R. Stromberg

School of Veterinary and Biomedical Sciences: Dissertations, Theses, and Student Research

Shiga toxin-producing Escherichia coli (STEC) are enteric pathogens of humans. Cattle serve as a reservoir and harbor STEC in their intestines. Intimin-positive STEC are referred to as enterohemorrhagic E.coli (EHEC). Seven serogroups (O26, O45, O103, O111, O121, O145, and O157) account for the majority of illness due to STEC and are hereafter referred to as STEC/EHEC-7. To improve detection, enrichment broths were compared for supporting growth of STEC-7 and STEC O104:H4 (STEC-8). In pure culture, STEC enriched in trypticase soy broth (TSB) had significantly greater growth compared to TSB containing antimicrobials. In fecal samples, E. coli broth enrichment yielded …

Cancer And Infection, Kathleen Hope Plummer

USF Tampa Graduate Theses and Dissertations

E. coli is the most frequently isolated Gram negative pathogen from bacteremia in cancer patients and is repeatedly recovered from many other extraintestinal illnesses. These infections are commonly endogenous in nature and interfere with the treatment of cancer resulting in increased healthcare costs, morbidity, and mortality rates. Cancer and the treatments related to cancer cause alterations in the microbiome of the gut and other organs. Despite this point, there is a serious lack of knowledge about the genetic types of E. coli infecting cancer patients. This gap results in vague prevention strategies and limited treatment options for cancer patients. Multi …

Effect Of Carvacrol-Loaded Nanoemulsions On A Bioluminescent Strain Of Escherichia Coli O157:H7, Clara Maria Vasquez Mejia

Open Access Theses

Nanoemulsions have been shown to be effective delivery vehicles for poorly water soluble natural antimicrobials. In this study the antimicrobial activity of carvacrol (5-isopropyl-2-methylphenol) nanoemulsions against a bioluminescent Escherichia coli O157:H7 was investigated using light emission as an indicator of cell viability. Different emulsifiers (Ultralec Lecithin and Tween 20), oils (Palm stearin and Coconut oil) and various carvacrol concentrations (0, 1, 2 and 2.5%) were evaluated. Bioluminescence was monitored in situ using a Hamamatsu (photo multiplier tube) sensor module integrated with a Programmable Logic Controller interfaced with a PC for data acquisition. Bioluminescence decreased rapidly with the addition of emulsions …

Comparison Of Two Separation Methods For Biological Particles: Field-Flow Fractionation (Fff) And Sucrose Density Gradients, Cody E. Garrison

OES Theses and Dissertations

Sucrose gradient centrifugation and Field-flow fractionation (FFF) are two different particle separation methods that overcome the problems of similar size microorganisms clumping together during standard filtration methods. FFF separates particles based on size and density via the parabolic velocity profile of laminar flow in a ribbon-like channel. The sucrose method separates particles via centrifugation in a density gradient. Both techniques worked well in separating eukaryotic from prokaryotic microbes, with the preferred method depending on the type and relative abundance of organisms to be separated. Minicells were separated from mother cells in transformed Escherichia coli cultures, heterotrophic flagellates (e.g., Diplonema papillatum …

Conservation Of The Che And Pho Systems In Escherichia Coli, Kimberly B. Zumbrennen, Dr. William R. Mccleary

Journal of Undergraduate Research

Bacteria live in a continuously changing environment. In order to survive, bacteria must monitor nutrients, acidity, temperature, osmolarity, etc. The use of two-component systems to sense and respond to these environmental changes is a common motif observed in a wide variety of organisms. A two-component system consists of two main proteins, a sensor and a response regulator(2). The sensor protein is a trans-membrane protein which monitors environmental fluctuations and activates the response regulator through phosphorylation. The response regulator contains two domains, the receiver module and the output domain. The receiver module is phosphorylated and dephosphorylated by the sensor protein. Communication …

Preventing Salmonella And Other Bacterial Contamination In Poultry Using A High-Adhering Strain Of Escherichia Coli, Jeffrey P. North, Dr. Ron Leavitt

Journal of Undergraduate Research

In 1928 Alexander Fleming revolutionized the world with the discovery of penicillin. Penicillin and the antibiotics discovered thereafter revolutionized the treatment of infectious diseases. Equipped with these miracle drugs, scientists thought the battle against pathogenic bacteria was won. However, bacteria began to evolve to meet the antibiotic challenge. As the 20th century progressed, more and more bacteria developed resistance to one or more antibiotics. This resistance is achieved through mutation or through horizontal gene transfer, when one bacterium transfers the gene to another bacterium.

Locating The High Adherence Genes On Pha-28, Jonathan Walter Dukes, Dr. Ronald Leavitt

Journal of Undergraduate Research

Bacterial high adherence is an important of research because of its effect on bacterial pathogenesis and intestinal colonization. In the laboratory of Dr. Ronald Leavitt, we have been studying the ability of certain bacteria to prevent the colonization of intestinal epithelial cells by other bacteria. Of particular interest to us are non-pathogenic Escherichia coli strains that have the ability to out compete and overrun colonies of Salmonella, as well as the colonies of the pathogenic E. coli strains that cause the turkey disease colibacillosis. Such E. coli have the potential to be used as an interfering agent against Salmonella and …

Phosphorylation Dependent Conformational Changes Of Phob In Escherichia Coli, Allison Sparks, Dr. William Mccleary

Journal of Undergraduate Research

All bacteria must have a way to control cell functions when their outside environment changes. E.coli uses a phosphate signaling pathway to accomplish this. In the bacterial cell membrane, there are proteins that take up phosphate from the environment, transports it into the cell, and triggers transcription of certain genes. The system is comprised of an ATP-dependent transport system which consists of a periplasmic substrate binding protein, PstS, three membrane proteins, PstC, PstA, and PstB, a regulator, PhoU, a transmembrane histidine kinase, PhoR, and a response regulator PhoB. In this system, the response regulator, PhoB is phosphorlated and then activates …

Determination And Analysis Of The Functionality And Required Amino Acid Residues Of Phou In Escherichia Coli, Michael Schurdell, Dr. William Mccleary

Journal of Undergraduate Research

While researching the required amino acid residues of PhoU, we discovered a small intergenic region located between pstA and pstB on the pstSCAB-phoU operon. Defining the role of this small intergenic region took precedence over my previously proposed research project. All of my research on this project was performed in conjunction with Garrett Woodbury and William McCleary. We presented our findings in a poster presentation at the Intermountain Branch of the American Society of Microbiology Annual Meeting on Saturday, March 18, 2006. A peer reviewed article containing the full details of our work was accepted for publication on December 1, …

Mscs-Like Mechanosensitive Channels In Plants And Microbes, Margaret E. Wilson, Grigory Maksaev, Elizabeth S. Haswell

Biology Faculty Research

The challenge of osmotic stress is something all living organisms must face as a result of environmental dynamics. Over the past three decades, innovative research and cooperation across disciplines have irrefutably established that cells utilize mechanically gated ion channels to release osmolytes and prevent cell lysis during hypoosmotic stress. Early electrophysiological analysis of the inner membrane of Escherichia coli identified the presence of three distinct mechanosensitive activities. The subsequent discoveries of the genes responsible for two of these activities, the mechanosensitive channels of large (MscL) and small (MscS) conductance, led to the identification of two diverse families of mechanosensitive channels. …

Understanding The Phou Protein’S Part In Phosphate Import By Synthesis Of Double Mutations In Escherichia Coli., Geoffrey R. Johnston, Dr. William Mccleary

Journal of Undergraduate Research

The purpose of our research was to gain a better understanding of what function the phoU protein serves in phosphate import by completely knocking out it’s functionality at two highly conserved and nearly identical sites on almost identical domains at a genetic level.

Isolation Of An Escherichia Coli High-Adherence Plasmid For Use In A Bacterial Interfering Agent, Christopher M. Runyan, Dr. Ronald Leavitt

Journal of Undergraduate Research

Colibacillosis is a disease found in poultry that results in various levels of morbidity and mortality. It is primarily caused by pathogenic strains of E. coli that inhabit the poults’ intestinal tracts. Antibiotics have frequently been used to protect the turkeys but with less and less effectiveness as the bacteria have begun to develop antibiotic resistance. Hence alternate ways are needed to combat pathogenic bacteria.

Phou Localization In Escherichia Coli, Austin Callison, Dr. William R. Mccleary

Journal of Undergraduate Research

Just as simpler systems are studied to better understand more complex systems, E. coli has long been a standard for biological experiments aimed at a more complete comprehension of the mechanisms that propitiate cell life. One of these vital mechanisms in E. coli is the uptake of phosphate (Pi) in varying levels of environmental phosphorous. Pi is a key element involved in cell membranes, DNA, RNA, and metabolism in E. coli. This uptake is accomplished by the cooperation of the PstSCAB2, PhoU, PhoR and PhoB proteins in phosphate poor environments1. PhoU acts as a negative regulator of the Pho operon …

Escherichia Coli Strain Diversity In Humans: Effects Of Sampling Effort And Methodology, Emily R. Neal

Master's Theses

Studies investigating Escherichia coli strain diversity and demographics in human hosts are frequently inconsistent regarding sampling effort and methodology while current strain typing methods are often expensive or laborious. To rectify these inconsistencies, sampling effort was investigated by comparing the diversity of 15-isolate collections to 100-isolate collections from 3 human subjects. Temporal variation in E. coli strain diversity was also studied by collecting 15 isolates once every 6 months. Additionally, strain identification and diversity collected by different sampling methods (fecal swabs vs. anal swabs collected at different times around defecation) were compared to identify any inherent biases in sampling method. …

Fecal Indicator Bacteria In Knitting Mill Creek With A Comparison Of Methods Used To Enumerate E. Coli, Manju Nagarajan

OES Theses and Dissertations

This research seeks to evaluate the impact of an urban, multi-use neighborhood on the bacteriological quality of water in Knitting Mill Creek, a blind arm of the Lafayette River, a sub-estuary of the lower Chesapeake Bay. A principal objective was to determine the effects of rainfall, a surrogate for run-off, on water-column concentrations of fecal-indicator bacteria (FIB). I collected water samples weekly (September 2009-December 2010) and biweekly (January 2010-December 2011) at a storm-sewer outfall and a nearby marina and quantified their E. coli and enterococci concentrations using Colilert-18 and Enterolert, most-probable-number assays. In addition, I took monthly samples for these …

An Investigation Of Bacterial Ribonucleases As An Antibiotic Target, Ashley Denise Frazier

Electronic Theses and Dissertations

Antibiotics have been commonly used in medical practice for over 40 years. However, the misuse and overuse of current antibiotics is thought to be the primary cause for the increase in antibiotic resistance.

Many current antibiotics target the bacterial ribosome. Antibiotics such as aminoglycosides and macrolides specifically target the 30S or 50S subunits to inhibit bacterial growth. During the assembly of the bacterial ribosome, ribosomal RNA of the 30S and 50S ribosomal subunits is processed by bacterial ribonucleases (RNases). RNases are also involved in the degradation and turnover of this RNA during times of stress, such as the presence of …

Toxin-Antitoxic Loci Vapbc-1 And Vapxd Contribute To Survival And Virulence In Nontypeable Haemophilus Influenzae, Dabin Ren, Anna N. Walker, Dayle A. Daines

Biological Sciences Faculty Publications

Background: Nontypeable Haemophilus influenzae (NTHi) is a significant human pathogen responsible for respiratory tract infections and the most common cause of recurrent otitis media. Type II toxin-antitoxin (TA) systems are genetic elements that code for a stable protein toxin and a labile antitoxin that are thought to be involved in metabolic regulation of bacteria by enabling a switch to a dormant state under stress conditions. The contribution to infection persistence of the NTHi TA loci vapBC-1 and vapXD was examined in this study.

Results: Deletions in vapBC-1, vapXD and vapBC-1 vapXD significantly decreased the survival of NTHi co-cultured with …

Modulation Of Alpha-Subunit Visit-Dg Sequence Residues Ser-347, Gly-351 And Thr-349 In The Catalytic Sites Of Escherichia Coli Atp Synthase., Laura Elaine Brudecki

Electronic Theses and Dissertations

Binding of inorganic phosphate (P_i) in ATP synthase catalytic sites is a crucial step for the synthesis of adenosine-5'-triphosphate (ATP). ATP is the fundamental means of cellular energy in almost every organism, and in order to gain insight into the regulation of ATP catalysis, critical amino acid residues responsible for binding P_i must be identified. Here, we investigate the role of highly conserved α-subunit VISIT-DG sequence residues αSer-347, αGly-351, and αThr-349 in P_i binding. Mutations αS347A/Q, αG351Q, αT349A/D/R, βR182A, and αT349R/βR182A were generated via site directed mutagenesis. Results from biochemical assays showed that αSer-347 is required …

Interaction Between Bacteriophage Dms3 And Host Crispr Region Inhibits Group Behaviors Of Pseudomonas Aeruginosa, Michael E. Zegans, Jeffrey C. Wagner, Kyle C. Cady, Daniel M. Murphy, John H. Hammond, George A. O'Toole

Dartmouth Scholarship

Bacteriophage infection has profound effects on bacterial biology. Clustered regular interspaced short palindromic repeats (CRISPRs) and cas (CRISPR-associated) genes are found in most archaea and many bacteria and have been reported to play a role in resistance to bacteriophage infection. We observed that lysogenic infection of Pseudomonas aeruginosa PA14 with bacteriophage DMS3 inhibits biofilm formation and swarming motility, both important bacterial group behaviors. This inhibition requires the CRISPR region in the host. Mutation or deletion of five of the six cas genes and one of the two CRISPRs in this region restored biofilm formation and swarming …

The Structural And Folding Characteristics Of The Plasmid-Encoded Toxin From Enteroaggregative Escherichia Coli, Patricia Scaglione

Electronic Theses and Dissertations

Plasmid-encoded toxin (Pet) from enteroaggregative Escherichia coli is a member of the autotransporter subfamily termed SPATE (serine protease autotransporters of Enterobacteriaceae). Autotransporters, which are the most common Gram-negative secreted virulence factors, contain three functional domains: an amino terminal leader sequence, a mature protein or passenger domain, and a carboxy-terminal β domain. The leader sequence targets the protein to the periplasmic space and the β domain then forms a β-barrel pore in the outer membrane of the bacterium which allows the passenger domain to enter the external milieu. In some cases the passenger domain is cleaved from the β-barrel at the …

A Serratia Marcescens Oxyr Homolog Mediates Surface Attachment And Biofilm Formation, Robert M. Q. Shanks, Nicholas A. Stella, Eric J. Kalivoda, Megan R. Doe

Dartmouth Scholarship

OxyR is a conserved bacterial transcription factor with a regulatory role in oxidative stress response. From a genetic screen for genes that modulate biofilm formation in the opportunistic pathogen Serratia marcescens, mutations in an oxyR homolog and predicted fimbria structural genes were identified. S. marcescens oxyR mutants were severely impaired in biofilm formation, in contrast to the hyperbiofilm phenotype exhibited by oxyR mutants of Escherichia coli and Burkholderia pseudomallei. Further analysis revealed that OxyR plays a role in the primary attachment of cells to a surface. Similar to what is observed in other bacterial species, S. marcescens OxyR …