Microbiology Commons^™

Characterization Of The Extracellular Proteome Of A Natural Microbial Community With An Integrated Mass Spectrometric / Bioinformatic Approach, Brian Keith Erickson

Doctoral Dissertations

Proteomics comprises the identification and characterization of the complete suite of expressed proteins in a given cell, organism or community. The coupling of high performance liquid chromatography (LC) with high throughput mass spectrometry (MS) has provided the foundation for current proteomic progression. The transition from proteomic analysis of a single cultivated microbe to that of natural microbial assemblages has required significant advancement in technology and has provided greater biological understanding of microbial community diversity and function.

To enhance the capabilities of a mass spectrometric based proteomic analysis, an integrated approach combining bioinformatics with analytical preparations and experimental data collection was …

Analysis Of Polymorphism In Human Cytomegalovirus (Hcmv) Chemokine, Vcxcl-1, And Its Role In Cellular Activation, Jinho Heo

Doctoral Dissertations

The human cytomegalovirus (HCMV) viral chemokine gene, UL146, shows a high degree of variability in clinical isolates. The UL146-produced viral chemokine, vCXCL-1, has homology to CXC chemokines and is predicted to be an immune modulator that may contribute to the pathogenesis of HCMV infections. In the analysis of clinical isolates from congenitally infected infants, we found 11 distinct vCXCL-1 clades. Although the four cysteine residues that create two disulfide bonds providing the essential structure for CXC chemokines,are conserved, the N-loop region, which is important for receptor binding and activation, was hypervariable. One clade also contained a modified glutamic acid-leucine-arginine (ELR) …

Study Of Population Diversity Of Toxoplasma Gondii, Debashree Majumdar

Masters Theses

Toxoplasma gondii, the causal agent of toxoplasmosis, is an important water and food borne protozoan parasite. T. gondii was previously shown to have a distinct clonal population structure composed of Type I, II and III lineages in North America and Europe. But more recent studies demonstrated high diversity in South America. In the present project we have conducted an intensive study of the population diversity of T. gondii and surveyed the extent of genetic variation among natural T. gondii isolates on a global scale in order to better understand the population dynamics and pathogenesis of this parasite. To this …

Mechanisms Of Antimicrobial Peptide Resistance In Campylobacter, Ky Van Hoang

Doctoral Dissertations

Campylobacter is the major bacterial cause of human gastroenteritis in the United States and other developed countries. Poultry are considered a main source of human Campylobacter infections. Thus, reduction of Campylobacter load in poultry is significant in food safety and public health. However, no effective measure is commercially available to prevent Campylobacter colonization in poultry to date. Antimicrobial peptides (AMPs) are short and bactericidal peptides widely present in intestine to limit bacterial infections. Recently, AMPs have been increasingly recognized as a novel class of antibiotics (peptide antibiotics) to control foodborne pathogens. Notably, several potent anti-Campylobacter bacteriocins, a group of AMPs …

T-Rfpred: A Nucleotide Sequence Size Prediction Tool For Microbial Community Description Based On Terminal-Restriction Fragment Length Polymorphism Chromatograms, Antonio Fernandez-Guerra, Alison Buchan, Xiaozhen Mou, Emilio O. Casamayor, Jose M. Gonzalez

Microbiology Publications and Other Works

Background

Terminal-Restriction Fragment Length Polymorphism (T-RFLP) is a technique used to analyze complex microbial communities. It allows for the quantification of unique or numerically dominant phylotypes in amplicon pools and it has been used primarily for comparisons between different communities. T-RFPred, Terminal-Restriction Fragment Prediction, was developed to identify and assign taxonomic information to chromatogram peaks of a T-RFLP fingerprint for a more comprehensive description of microbial communities. The program estimates the expected fragment size of representative 16S rRNA gene sequences (either from a complementary clone library or from public databases) for a given primer and restriction enzyme(s) and provides candidate …

An Analysis Of Global Gene Expression Resulting From Exposure To Energetic Materials, Vernon L. Mcintosh Jr.

Doctoral Dissertations

AN ANALYSIS OF GLOBAL GENE EXPRESSION RESULTING FROM EXPOSURE TO ENERGETIC MATERIALS

A Dissertation

Presented for the Doctor of Philosophy Degree

University of Tennessee, Knoxville

VERNON LASHAWN MCINTOSH JR.

August 2010

Dedication

This dissertation is dedicated to my family. My mother and father Debra and Vernon McIntosh instilled in me the respect for academic excellence and the drive maximize my potential. Early on, my younger brother Kyle started showing signs of a shared interest in biology thus my desire to be a positive role model for him kept me motivated. Last but certainly not least, my loving wife and best …

5’-Proximal Cis-Acting Rna Signals For Coronavirus Genome Replication, Bo-Jhih Guan

Doctoral Dissertations

RNA sequences and higher-order structures in the 5’ and 3’ untranslated regions (UTRs) of positive-strand RNA viruses are known to function as cis-acting elements for translation, replication, and transcription. In coronaviruses, these are best characterized in the group 2a bovine coronavirus (BCoV) and mouse hepatitis virus (MHV), yet their precise mechanistic features are largely undefined. Here, we use a reverse genetics system in MHV to exploit the ~30% nt sequence divergence between BCoV and MHV to establish structure/function relationships of 5’ UTR cis-replication elements. It had been previously shown that a precise replacement of the 391-nt MHV 3’ UTR with …

Methicillin Resistance In Staphylococcus Pseudintermedius, Chad Christopher Black

Doctoral Dissertations

Staphylococcus pseudintermedius affecting dogs is analogous to S. aureus on humans, acting as both normal flora and opportunistic pathogen. Methicillin resistance in S. pseudintermedius is recent, with the first documented occurrence of an isolate bearing the methicillin resistance gene, mecA, in 1999. This gene encodes penicillin binding protein 2a, which renders all beta-lactam drugs ineffective and functions as a “gateway” antibiotic resistance determinant. In the presence of ineffective antibiotics, opportunities for mutational events and acquisition of mobile genetic elements increase as microbial densities increase, often leading to multi-drug resistance. Methicillin-resistant S. pseudintermedius (MRSP) infections have become increasingly common. For …

Characterizing Cell-Cell And Cell-Surface Interactions In The Rhizobacterium Azospirillum Brasilense, Calvin Shay Green

Masters Theses

Microaerophilic and chemotaxic diazotrophs, azospirilla are found in close association with certain cereals such as durum wheat and maize and are active in enriching these ecological niches with the macronutrient nitrogen as ammonia. Regarded as highly pleomorphic, Azospirillum spp. are highly motile, using either a single polar flagellum when grown in liquid environments or peritrichous lateral flagella in viscous environments. Additionally, azospirilla are able to adhere onto surfaces as a biological film or aggregate cell-to-cell as nonproliferating flocculi, and these two processes having been suggested as positively affecting the survival and dispersal of the bacteria in the soil. Even though …

Influenza-Specific B Cell Responses In Hla-Dr1 Transgenic Mice, Lifang Huan

Masters Theses

HLA-DR1 transgenic (DR1 Tg) mice provide a model for evaluating the breadth and specificity of CD4 T cell responses that may develop in humans following influenza infection or vaccination. Recent studies identified a tremendously broad HLA-DR1-restricted CD4 T cell responses in DR1 Tg mice infected intranasally with influenza A/New Caledonia/20/99 (NC). In this study, our goals were to characterize B cell responses after NC infection in DR1 Tg mice and establish the correlation between B cell responses and CD4 T cell responses in this system. Influenza-specific B cell responses following virus administration were analyzed in DR1 Tg mice and in …

Cattle Access Affects Periphyton Community Structure In Tennessee Farm Ponds., Robert Gerald Middleton

Masters Theses

Cattle farming is vital to the economy of the United States. Frequently, cattle are given access to ponds and streams for water. The relative impacts of cattle access in natural water sources on the periphyton community have been rarely investigated. Periphyton is the basis of the aquatic food web, and community composition can serve as a bioindicator of pollution. Thus, my objectives were to quantify the effects of cattle access in aquatic lentic systems on periphyton community structure and biovolume, identify taxa that were associated with cattle access, and identify abiotic mechanisms that might be driving assemblage changes. I conducted …

Campylobacter Pathogenesis And Subunit Vaccine Development, Ximin Zeng

Doctoral Dissertations

Campylobacter jejuni is the leading bacterial cause of human gastroenteritis in the United States. Increasing resistance of Campylobacter to clinical antibiotics raises an urgent need for novel strategies to prevent and control infections in humans and animal reservoirs, which necessitates a better understanding of Campylobacter pathogenesis. We hypothesize that multidrug efflux pump CmeABC and ferric enterobactin (FeEnt) iron acquisition systems, which play a critical role in Campylobacter pathogenesis, are novel targets for developing effective measures against Campylobacter. To test this, the molecular, antigenic, functional, and protective characteristics of two outer membrane proteins, CmeC (an essential component of CmeABC drug efflux …

Naïve T Cells Re-Distribute To The Lungs Of Selectin Ligand Deficient Mice, Thandi M. Onami, John R. Harp

Microbiology Publications and Other Works

BACKGROUND: Selectin mediated tethering represents one of the earliest steps in T cell extravasation into lymph nodes via high endothelial venules and is dependent on the biosynthesis of sialyl Lewis X (sLe(x)) ligands by several glycosyltransferases, including two fucosyltransferases, fucosyltransferase-IV and -VII. Selectin mediated binding also plays a key role in T cell entry to inflamed organs.

METHODOLOGY/PRINCIPAL FINDINGS: To understand how loss of selectin ligands (sLe(x)) influences T cell migration to the lung, we examined fucosyltransferase-IV and -VII double knockout (FtDKO) mice. We discovered that FtDKO mice showed significant increases (approximately 5-fold) in numbers of naïve T cells in …

Coupling Metabolism And Chemotaxis-Dependent Behaviours By Energy Taxis Receptors, Gladys Alexandre

Microbiology Publications and Other Works

Bacteria have evolved the ability to monitor changes in various physico-chemical parameters and to adapt their physiology and metabolism by implementing appropriate cellular responses to these changes. Energy taxis is a metabolism-dependent form of taxis and is the directed movement of motile bacteria in gradients of physico-chemical parameters that affect metabolism. Energy taxis has been described in diverse bacterial species and several dedicated energy sensors have been identified. The molecular mechanism of energy taxis has not been studied in as much detail as chemotaxis, but experimental evidence indicates that this behaviour differs from metabolism-independent taxis only by the presence of …

Use Of Proteomics Tools To Investigate Protein Expression In Azospirillum Brasilense, Gurusahai K. Khalsa-Moyers

Doctoral Dissertations

Mass spectrometry based proteomics has emerged as a powerful methodology for investigating protein expression. “Bottom up” techniques in which proteins are first digested, and resulting peptides separated via multi-dimensional chromatography then analyzed via mass spectrometry provide a wide depth of coverage of expressed proteomes. This technique has been successfully and extensively used to survey protein expression (expression proteomics) and also to investigate proteins and their associated interacting partners in order to ascertain function of unknown proteins (functional proteomics). Azospirillum brasilense is a free-living diazotrophic soil bacteria, with world-wide significance as a plant-growth promoting bacteria. Living within the rhizosphere of cereal …

Agricultural Soil Bacteria; A Study Of Collection, Cultivation, And Lysogeny, Katherine Elizabeth Sides

Masters Theses

The aim of this research project was to test new collection and cultivation techniques that may increase the range of cultivable diversity of soil bacteria. Fortified BioSep beads were employed in situ to capture soil bacteria, and the success of the beads was analyzed using Phylochip microarray analysis. In the cultivation phase, three different media substrates and increased incubation period were evaluated for the ability to select novel or rare bacteria. Over 700 agricultural soil bacterial isolates were classified, including a rare Gemmatimonadetes sp., a rare Verrucomicrobia sp., several Acidobacteria sp., and many novel isolates. Land management, media, and incubation …

The Microscope (2010-2011), Department Of Microbiology

The Microscope

No abstract provided.

The Construction And Analysis Of Marker Gene Libraries, S.M. Short, F. Chen, Steven Wilhelm

Microbiology Publications and Other Works

Marker genes for viruses are typically amplified from aquatic samples to determine whether specific viruses are present in the sample, or to examine the diversity of a group of related viruses. In this chapter, we will provide an overview of common methods used to amplify, clone, sequence, and analyze virus marker genes, and will focus our discussion on viruses infecting algae, bacteria, and heterotrophic flagellates. Within this chapter, we endeavor to highlight critical aspects and components of these methods. To this end, instead of providing a detailed experimental protocol for each of the steps involved in examining virus marker gene …

Determining Rates Of Virus Production In Aquatic Systems By The Virus Reduction Approach,, M.G. Weinbauer, J.M. Rowe, Steven Wilhelm

Microbiology Publications and Other Works

The reduction approach to assess virus production and the prokaryotic mortality by viral lysis stops new infection by reducing total virus abundance (and thus virus–host contacts). This allows for easy enumeration of viruses that originate from lysis of already infected cells due to the decreased abundance of free virus particles. This reoccurrence can be quantified and used to assess production and cell lysis rates. Several modifications of the method are presented and compared. The approaches have great potential for elucidating trends in virus production rates as well as for making generalized estimates of the quantitative effects of viruses on marine …

Spatial Structure And Activity Of Sedimentary Microbial Communities Underlying A Beggiatoa Spp. Mat In A Gulf Of Mexico Hydrocarbon Seep, Karen Lloyd, Daniel B. Albert, Jennifer F. Biddle, Jeffrey P. Chanton, Oscar Pizarro, Andreas Teske

Microbiology Publications and Other Works

Background

Subsurface fluids from deep-sea hydrocarbon seeps undergo methane- and sulfur-cycling microbial transformations near the sediment surface. Hydrocarbon seep habitats are naturally patchy, with a mosaic of active seep sediments and non-seep sediments. Microbial community shifts and changing activity patterns on small spatial scales from seep to non-seep sediment remain to be examined in a comprehensive habitat study.

Methodology/Principal Findings

We conducted a transect of biogeochemical measurements and gene expression related to methane- and sulfur-cycling at different sediment depths across a broad Beggiatoa spp. mat at Mississippi Canyon 118 (MC118) in the Gulf of Mexico. High process rates within the …

Autonomous Bioluminescent Expression Of The Bacterial Luciferase Gene Cassette (Lux) In A Mammalian Cell Line, Dan M. Close, Stacey S. Patterson, Steven Ripp, Seung J. Baek, John Sanseverino, Gary S. Sayler

Microbiology Publications and Other Works

Background

The bacterial luciferase (lux) gene cassette consists of five genes (luxCDABE) whose protein products synergistically generate bioluminescent light signals exclusive of supplementary substrate additions or exogenous manipulations. Historically expressible only in prokaryotes, the lux operon was re-synthesized through a process of multi-bicistronic, codon-optimization to demonstrate for the first time self-directed bioluminescence emission in a mammalian HEK293 cell line in vitro and in vivo.

Methodology/Principal Findings

Autonomous in vitro light production was shown to be 12-fold greater than the observable background associated with untransfected control cells. The availability of reduced riboflavin phosphate (FMNH2) was identified as the limiting bioluminescence substrate …