Bacteriology Commons^™

Enhanced Expression Of Codon Optimized Mycobacterium Avium Subsp. Paratuberculosis Antigens In Lactobacillus Salivarius, Christopher D. Johnston, John P. Bannatine, Rodney Govender, Lorraine Endersen, Daniel Pletzer, Helge Weingart, Aidan Coffey, Jim O'Mahony, Roy D. Sleator

Department of Biological Sciences Publications

It is well documented that open reading frames containing high GC content show poor expression in A+T rich hosts. Specifically, G+C-rich codon usage is a limiting factor in heterologous expression of Mycobacterium avium subsp. paratuberculosis (MAP) proteins using Lactobacillus salivarius. However, re-engineering opening reading frames through synonymous substitutions can offset codon bias and greatly enhance MAP protein production in this host. In this report, we demonstrate that codon-usage manipulation of MAP2121c can enhance the heterologous expression of the major membrane protein (MMP), analogous to the form in which it is produced natively by MAP bacilli. When heterologously over-expressed, antigenic determinants …

Characterizing The Response Of Multidrug-Resistant Klebsiella Pneumoniae Species To The Application Of A Phage Cocktail, Steven Liu

Symposium

Project Summary: The application of bacteriophages to treat bacterial infections is known as phage therapy, which takes advantage of bacteriophage’s natural ability to infect and lyse bacterial hosts. Phages have been shaped by billions of years of evolution to be highly specialized deliverers of bactericidal agents to the cytoplasm of their target bacteria. Ever since discovery of bacteriophages in 1915, phage therapy was recognized as a potentially powerful tool for eliminating bacterial infections. The effectiveness of phage therapy can be increased by creating a mixture of multiple phages to target a wider variety of bacterial strains. Furthermore, phage therapy has …

Characterization Of Ftsa-Ftsn Interaction During Escherichia Coli Cell Division, [email protected] K. Busiek

Dissertations and Theses (Open Access)

Division of a bacterial cell into two equal daughter cells requires precise assembly and constriction of the division machinery, or divisome. The Escherichia coli divisome includes nearly a dozen essential cell division proteins that assemble at midcell between segregating sister chromosomes. FtsZ, a homolog of eukaryotic tubulin, is the first essential cell division protein to localize at midcell where it polymerizes into a ring-shaped scaffold (Z ring). Establishment of the Z ring is required for recruitment of downstream cell division proteins including FtsA, a cytoplasmic protein that tethers the Z ring to the inner membrane. Following localization of FtsA and …

Transcriptome Analysis Of Listeria Monocytogenes Exposed To Biocide Stress Reveals A Multi-System Response Involving Cell Wall Synthesis, Sugar Uptake, And Motility, Aidan Casey, Edward M. Fox, Stephan Schmitz-Esser, Aidan Coffey, Olivia Mcauliffe, Kieran Jordan

Department of Biological Sciences Publications

Listeria monocytogenes is a virulent food-borne pathogen most often associated with the consumption of “ready-to-eat” foods. The organism is a common contaminant of food processing plants where it may persist for extended periods of time. A commonly used approach for the control of Listeria monocytogenes in the processing environment is the application of biocides such as quaternary ammonium compounds. In this study, the transcriptomic response of a persistent strain of L. monocytogenes (strain 6179) on exposure to a sub-lethal concentration of the quaternary ammonium compound benzethonium chloride (BZT) was assessed. Using RNA-Seq, gene expression levels were quantified by sequencing …

Phages Of Non-Diary Lactococci: Isolation And Characterization Of Phi L47, A Phage Infecting The Grass Isolate Lactococcus Lactis Ssp Cremoris Dpc6860, Daniel Cavanagh, Caitríona M. Guinane, Horst Neve, Aidan Coffey, R. Paul Ross, Gerald F. Fitzgerald, Olivia Mcauliffe

Department of Biological Sciences Publications

Lactococci isolated from non-dairy sources have been found to possess enhanced metabolic activity when compared to dairy strains. These capabilities may be harnessed through the use of these strains as starter or adjunct cultures to produce more diverse flavor profiles in cheese and other dairy products. To understand the interactions between these organisms and the phages that infect them, a number of phages were isolated against lactococcal strains of non-dairy origin. One such phage, ΦL47, was isolated from a sewage sample using the grass isolate L. lactis ssp. cremoris DPC6860 as a host. Visualization of phage virions by transmission electron …

Listeria Monocytogenes Can Utilize Both M Cell Transcytosis And Inla-Mediated Uptake To Cross The Epithelial Barrier Of The Intestine During An Oral Infection Model Of Listeriosis, Hilary Denney

Theses and Dissertations--Medical Sciences

The invasive pathways, InlA- and InB-mediated uptake and M cell transcytosis, that Listeria monocytogenes uses to invade the intestine have mainly been studied using infection models that do not truly replicate what occurs during a natural infection. Recently, our lab has developed an oral infection model that is more physiolocally relevant to what occurs during food borne listeriosis. We have sought to evaluate the relative roles of the previously defined invasive pathways, in our oral model of infection. We have done this by utilizing an InlAmCG Lm strain that is able to bind murine E-cadherin, knockout Lm strains, ΔinlA Lm …

A Prospective Study Of Periodontal Disease And Risk Of Gastric And Duodenal Ulcer In Male Health Professionals, Matthew R. Boylan, Hamed Khalili, Edward S. Huang, Dominique S. Michaud, Jacques Izard, Kaumudi J. Joshipura, Andrew T. Chan

Department of Food Science and Technology: Faculty Publications

Periodontal disease is characterized by microbial shifts within the oral cavity and chronic inflammation of the gingival tissue, and can lead to tooth loss. Several studies support an association between periodontal disease and risk of chronic systemic conditions, including vascular disease, chronic obstructive pulmonary disease, diabetes mellitus, and cancer, including gastric adenocarcinoma. These associations may be mediated by systemic inflammation, the immune response to periodontal infection, or direct invasion by pathogens. Previous studies of periodontal disease and risk of peptic ulcer have been limited by cross-sectional design, diagnoses that were not confirmed by medical record review, and a lack of …

Regulation Of Hyaluronate Lyase (Hyla) Expression By Regr A Transcriptional Repressor In Streptococcus Pyogenes, Alexis A. Kordis

Biological Sciences Theses & Dissertations

Streptococcus pyogenes, group A streptococci (GAS), causes diseases ranging from asymptomatic to life threatening. Some strains of S. pyogenes produce extracellular hyaluronate lyase (HylA), a potential virulence factor. HylA is an enzyme that degrades hyaluronic acid, which is found in the extracellular matrix of human tissues. The breakdown of the host tissue contributes to the spread of infection. The hylA gene is not constitutively expressed in vitro, which implies regulation.

A proposed regulator of hyaluronate lyase expression is RegR, a LacI/GalR like protein. A vector containing disrupted regR was electro-transformed into S. pyogenes M-type 22 strain 10403. Growth characteristics …

Composition Of The Adult Digestive Tract Bacterial Microbiome Based On Seven Mouth Surfaces, Tonsils, Throat And Stool Samples, Nicola Segata, Susan Kinder Haake, Peter Mannon, Katherine P. Lemon, Levi Waldron, Dirk Gevers, Curtis Huttenhower, Jacques Izard

Department of Food Science and Technology: Faculty Publications

Background: To understand the relationship between our bacterial microbiome and health, it is essential to define the microbiome in the absence of disease. The digestive tract includes diverse habitats and hosts the human body’s greatest bacterial density. We describe the bacterial community composition of ten digestive tract sites from more than 200 normal adults enrolled in the Human Microbiome Project, and metagenomically determined metabolic potentials of four representative sites.

Results: The microbiota of these diverse habitats formed four groups based on similar community compositions: buccal mucosa, keratinized gingiva, hard palate; saliva, tongue, tonsils, throat; sub- and supra-gingival plaques; …

Efficient And Robust Rna-Seq Process For Cultured Bacteria And Complex Community Transcriptomes, Georgia Giannoukos, Dawn M. Ciulla, Katherine Huang, Brian J. Haas, Jacques Izard, Joshua Z. Levin, Jonathan Livny, Ashlee M. Earl, Dirk Gevers, Doyle V. Ward, Chad Nusbaum, Bruce W. Birren, Andreas Gnirke

Department of Food Science and Technology: Faculty Publications

We have developed a process for transcriptome analysis of bacterial communities that accommodates both intact and fragmented starting RNA and combines efficient rRNA removal with strand-specific RNA-seq. We applied this approach to an RNA mixture derived from three diverse cultured bacterial species and to RNA isolated from clinical stool samples. The resulting expression profiles were highly reproducible, enriched up to 40-fold for non-rRNA transcripts, and correlated well with profiles representing undepleted total RNA.

Staphylococcus Aureus Virulence Factors Synthesis Is Controlled By Central Metabolism, Yefei Zhu

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

Staphylococcus aureus is a versatile pathogen that can survive in diverse host environments. This versatility depends on its ability to sense nutrients and respond by modulating gene expression, including the synthesis of virulence determinants. In addition to its ability to synthesize virulence factors, the capacity of S. aureus to form biofilms is an important mediator of virulence in certain infections. Biofilms are a complex aggregation of bacteria commonly encapsulated by an adhesive exopolysaccharide matrix (polysaccharide intercellular adhesin; PIA). To study S. aureus biofilm formation, we assessed the metabolic requirements of S. aureus growing in a biofilm and …

Killing Of Treponema Denticola By Mouse Peritoneal Macrophages, P. Gaibani, C. Vocale, S. Ambretti, F. Cavrini, Jacques Izard, L. Miragliotta, M. T. Pellegrino, V. Sambri

Department of Food Science and Technology: Faculty Publications

Treponema denticola has been identified as an important cause of periodontal disease and hypothesized to be involved in extra-oral infections. The objective of this study was to investigate the role of T. denticola cell length and motility during mouse peritoneal macrophages in vitro uptake. Macrophages, incubated under aerobic and anaerobic conditions, produced a similar amount of TNF-α when stimulated with Escherichia coli LPS. The uptake of FlgE- and CfpA-deficient mutants of T. denticola was significantly increased compared with the wild-type strain, due to cell size or lack of motility. Opsonization with specific antibodies considerably improved the treponemes’ uptake. …

Native Cellular Architecture Of Treponema Denticola Revealed By Cryo-Electron Tomography, Jacques Izard, Chyong-Ere Hsieh, Ronald J. Limberger, Carmen A. Mannella, Michael Marko

Department of Food Science and Technology: Faculty Publications

Using cryo-electron tomography, we are developing a refined description of native cellular structures in the pathogenic spirochete Treponema denticola. Tightly organized bundles of periplasmic flagella were readily observed in intact plunge-frozen cells. The periplasmic space was measured in both wild-type and aflagellate strains, and found to widen by less than the diameter of flagella when the latter are present. This suggests that a structural change occurs in the peptidoglycan layer to accommodate the presence of the flagella. In dividing cells, the flagellar filaments were found to bridge the cytoplasmic cylinder constriction site. Cytoplasmic filaments, adjacent to the inner membrane, …

Cryo-Electron Tomography Elucidates The Molecular Architecture Of Treponema Pallidum, The Syphilis Spirochete, Jacques Izard, Christian Renken, Chyong-Ere Hsieh, Daniel C. Desrosiers, Star Dunham-Ems, Carson La Vake, Linda L. Gebhardt, Ronald J. Limberger, David L. Cox, Michael Marko, Justin D. Radolf

Department of Food Science and Technology: Faculty Publications

Cryo-electron tomography (CET) was used to examine the native cellular organization of Treponema pallidum, the syphilis spirochete. T. pallidum cells appeared to form flat waves, did not contain an outer coat and, except for bulges over the basal bodies and widening in the vicinity of flagellar filaments, displayed a uniform periplasmic space. Although the outer membrane (OM) generally was smooth in contour, OM extrusions and blebs frequently were observed, highlighting the structure’s fluidity and lack of attachment to underlying periplasmic constituents. Cytoplasmic filaments converged from their attachment points opposite the basal bodies to form arrays that ran roughly …

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, …

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 …

Effect Of Short Exposure Of Naoci/Edta On E. Faecalis In Extracted Human Teeth, Rylan N. Gustafson

Loma Linda University Electronic Theses, Dissertations & Projects

The purpose of this project was to study the antibacterial effect of the short exposure of 5.25% NaOCl and 17% EDTA on eight strains of Enterococcus faecalis in extracted human teeth. The crowns of 240 single rooted teeth were removed and the canals were instrumented to ISO 0.36. The canals were irrigated with 5.25% NaOCl and 17% EDTA, autoclaved and separated into eight groups of 30 teeth consisting of 20 experimental teeth, 5 positive and 5 negative controls. Each group was inoculated with one of eight different strains of E. faecalis in BHl broth and incubated for four weeks. The …

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 …

Antibiotic Resistant And Coliform Bacteria In The Ohio River; 2002 To 2004, Lisa Marie Smith

Theses, Dissertations and Capstones

During 2002 and 2003 samples, collected every five miles along the Ohio River, were analyzed for total cultivable bacteria, ciprofloxacin resistant bacteria, erythromycin resistant bacteria, tetracycline resistant bacteria, and fecal coliforms. During 2004 samples were analyzed for total cultivable bacteria, ciprofloxacin resistant bacteria, sulfamethizole resistant bacteria, tetracycline resistant bacteria, Virginiamycin resistant bacteria, total coliforms, and Escherichia coli. The objectives of this study were to systematically collect data on fecal coliforms, E. coli, and antibiotic resistant bacteria in the Ohio River and its major tributaries; to determine if antibiotic resistance populations are correlated to each other or to coliforms; and …

Detection Of Bacteria To Dental Tissue By Low-Light Imaging, G-Hong Robert Hsu

Loma Linda University Electronic Theses, Dissertations & Projects

Bacteria have been shown to play an important role in the development of pulp and periradicular disease. The source of the infective bacteria is most often the oral cavity. Previous studies have detected bacteria in teeth that are structurally intact that seemingly have no communication intraorally. Anachoresis has been proposed as a mechanism for localizing circulating bacteria to such teeth. In several previous bacteriologic anachoresis studies, bacteria have been found to localize in inflamed dental tissues. The criticism most commonly associated with these investigations has been the potential contribution of contaminant bacteria resulting in false positive findings.

Low-light image analysis …

Characterization Of Spy1600 A Putative Hyaluronidase Gene In Group A Streptococci, Karin M. Berling

Biological Sciences Theses & Dissertations

Group A Streptococci (GAS), also known as Streptococcus pyogenes, can cause a variety of human diseases ranging from asymptomatic to life threatening. Exactly how a single type of organism is able to inflict such a multitude of diseases remains to be fully understood. One possibility includes the large number of secreted virulence factors expressed by the organism. The recent sequencing of three streptococcal genomes has indicated the existence of several previously unknown genes, some of which may encode possible virulence factors. Among these is Spy1600, which based on its sequence similarities has been proposed to encode a hyaluronidase, a …

Development Of An Electrotransformation Technique For Streptococcus Iniae And Preliminary Characterization Of The Hemolysin Associated With This Bacterium, Kimberly Ann Hahn

Biological Sciences Theses & Dissertations

Streptococcus iniae is a gram-positive organism responsible for causing disease in both freshwater as well as saltwater fish. Every year fisheries lose fish by the tons due to diseases caused by this organism. In 1991, the first reported human case of disease associated with this organism was described. Since this initial case, there have been numerous other reported cases of S. iniae infections. This organism is a catalase negative, facultatively anaerobic organism that produces a capsule and when plated onto blood agar demonstrates β-hemolytic activity.

In this study, the relationship between the hemolysin produced by S. iniae and the lactate …

Effect Of Magnesium On The Production Of Scna And Scnr By Streptococcus Pyogenes, Strain Ff22, Laura E. Quinn

Biological Sciences Theses & Dissertations

The lantibiotic SA-FF22, produced by Streptococcus pyogenes strain FF22, was the first described antimicrobial peptide produced by a Streptococcus species. S. pyogenes is a group A Streptococcus responsible for such human illness as strep-throat, rheumatic fever, necrotizing fasciitis, endocarditis, and meningitis. Previous studies of SA-FF22 have shown that antimicrobial activity is lost in the presence of lmM magnesium. We hypothesize that the lack of SA-FF22 activity in the presence of magnesium is due to an absence of transcription of scnA, the gene encoding SA-FF22. The lack of transcriptional activation of scnA may be due to an absence of ScnR, a …

Investigations Into The Innate Resistance Of The Tick, Dermacentor Variabilis, When Challenged With The Bacterium, Escherichia Coli, Shane Michael Ceraul

Biological Sciences Theses & Dissertations

In addition to the soluble response, many invertebrates control bacterial infections by means of phagocytosis or melanotic encapsulation. In some insects, Escherichia coli growth is reported to be inhibited by aggregation/encapsulation. Soluble and phagocytic responses to bacterial challenge have been reported in ticks, but evidence of an aggregation/encapsulation response was reported only for inanimate (araldite) implants. This study was done to determine how ticks control infection by E. coli. Ticks were challenged by direct inoculation of bacteria into the hemocoel cavity. Using plate counts, no viable E. coli were detected I hour post-inoculation. A direct fluorescence assay (DF A) …

Cytoplasmic Filament-Deficient Mutant Of Treponema Denticola Has Pleiotropic Defects, Jacques Izard, William A. Samsonoff, Ronald J. Limberger

Department of Food Science and Technology: Faculty Publications

In Treponema denticola, a ribbon-like structure of cytoplasmic filaments spans the cytoplasm at all stages of the cell division process. Insertional inactivation was used as a first step to determine the function of the cytoplasmic filaments. A suicide plasmid was constructed that contained part of cfpA and a nonpolar erythromycin resistance cassette (ermF and ermAM) inserted near the beginning of the gene. The plasmid was electroporated into T. denticola, and double-crossover recombinants which had the chromosomal copy of cfpA insertionally inactivated were selected. Immunoblotting and electron microscopy confirmed the lack of cytoplasmic filaments. The mutant was further …

Molecular Evolution Of Insecticidal Spore-Forming Bacteria, John Pool

Honors Theses

Molecular methods are increasingly being used to determine the phylogeny of microorganisms. This research was intended to determine phylogenetic relationships for bacteria of the species Bacillus thuringiensis and other members of the Bacillus cereus group. Each strain was analyzed by its sasp-B gene sequence to determine its species classification and relation to other strains studied. Results of this study indicated that according to the sasp-B gene tree, the species Bacillus thuringiensis is a paraphyletic with respect to both Bacillus cereus and Bacillus anthracis. Some unexpected results and implications for species designations are also discussed.

An Extracellular Protein Produced By Staphylococcus Hominis Strain Αm With Antimicrobial Activity Against Mycobacterium Spp, Sandra May Jacobsen

Biological Sciences Theses & Dissertations

The inhibitory activity exhibited by Staphylococcus hominis strain αM toward Mycobacterium species was examined. These studies included determining the conditions that maximized inhibitory agent production, analyzing characteristics of the agent, determining the time of agent production, ascertaining plasmid possession and the role of plasmids in the production of this inhibitory agent, and ascertaining the antimicrobial activity of other strains of S. hominis. On solid and in liquid media, S. hominis strain αM exhibits a unique inhibitory activity toward Mycobacterium species that is not demonstrated by Staphylococcus epidermidis or other S. hominis strains. Maximum agent production and recovery was …

Insertional Inactivation Of Treponema Denticola Tap1 Results In A Nonmotile Mutant With Elongated Flagellar Hooks, Ronald J. Limberger, Linda L. Slivienski, Jacques Izard, William A. Samsonoff

Department of Food Science and Technology: Faculty Publications

The treponemal fla operon is comprised of numerous motility-related genes; however, the initial gene of this operon, tap1, has no known function. A recently developed system to generate specific mutants in Treponema denticola was utilized to determine if Tap1 was essential for motility. T. denticola tap1 and flanking DNA were identified, cloned, and sequenced, and a suicide plasmid that contained tap1 interrupted with an erythromycin resistance cassette (ermF and ermAM) was constructed. Because of potential polar effects from this cassette, a second plasmid that contained tap1 interrupted with a modified erythromycin resistance cassette that lacked the putative ermF transcription …

Characterization Of Metal-Regulated Genes In Pseudomonas Fluorescens, Melanie Pearson

Honors Theses

One side effect of today's industrialized world is increased levels of heavy metals in the environment. Many of these metals are necessary for biological function as trace elements, but at higher concentrations are toxic. Other metals, such as cadmium, are not beneficial at any level, and have only deleterious effects on living organisms. Cadmium is primarily thought to interfere with normal biological function of proteins. Human exposure to cadmium appears to primarily damage the kidneys, but may also affect the liver, lungs, immune system, and central nervous system. Bacteria, however, have developed several methods for handling toxic heavy metals: cation …

Cellular Responses In Escherichia Coli To Lethal And Sublethal Doses Of Ozone, Indira Ruth Komanapalli

Loma Linda University Electronic Theses, Dissertations & Projects

Ozone is a major component of photochemical smog. High levels of this pollutant, sufficient to affect human health are found in many urban areas worldwide. Though limited studies in humans are supported by extensive findings from animal experiments, a difficulty in interpreting the results of these experiments has lead to an ambiguity on the biochemical mechanism of ozone toxicity. To elucidate the mechanism by which ozone causes cell damage and eventual cell death we conducted a comprehensive study using Escherichia coli K-12 as a model.

Studies on the comparative inactivation of bacteriophage lambda (λ), Escherichia coli, and Candida albicans …