Microbiology Commons^™

The Development Of New Molecular Tools To Detect Mycobacterium Avium Subspecies Paratuberculosis From A Bovine Faecal Environment By Real-Time Pcr, Marcel De Kruijf

Theses

The slow-growing intracellular pathogen Mycobacterium avium subspecies paratuberculosis (MAP) causes Johne’s disease in domesticated and wild ruminants. Johne’s disease inflicts severe annual economic losses to the dairy industry worldwide. This thesis is divided into five chapters that consist of a literature review' chapter and four experimental chapters focusing on MAP strain genotyping and the development of novel molecular MAP detection tools using real-time PCR. Chapter 1 describes published information regarding MAP characteristics, Johne’s disease, MAP genomes, available MAP detection methods, MAP epidemiology and genotyping.

To efficiently control and monitor MAP infection, molecular t>ping of MAP strains provides extensive knowledge …

Molecular Biology Of A Surface-Exposed Protein Family Of Bartonella Bacilliformis, Hannah Fay

Undergraduate Theses, Professional Papers, and Capstone Artifacts

Bartonella bacilliformis is a gram negative alpha-proteobacterium native to the Andes of South America. Bartonella causes Carrion’s disease, a potentially life threatening disease transmitted by the sand fly. It has a mortality rate of 88% when untreated and 10% when treated. Diagnostics and control measures for the disease are underdeveloped and no vaccine is available. Recent outbreaks indicate that the range of the pathogen is increasing with nearly 1.7 million people in western South America at risk. Little is known about the epidemiology and pathogenesis of B. bacilliformis. Recent research done by UC San Diego found a paralogous gene …

Vitamin D3 And Suppressor Of Cytokine Signaling Proteins Reduces Pro-Inflammatory Cytokines In An Alzheimer’S Disease Like-Model Consisting Of Microglial And Neuronal Co-Cultures, Alexander Evdokiou

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This study examined the inflammatory effects of amyloid-ß (Aß42) in a microglial-neuronal co-culture system and determined whether 1a, 25-dihydroxyvitamin D3 (1,25-(OH)2D3) along with suppressor of cytokine signaling (SOCS)1 and SOCS 3 mimetics would attenuate the inflammatory response to Aß42. This culture system, when seeded with Aß42, serves as an in vitro model for Alzheimer’s disease (AD). In a neuronal-microglia co-culture, Aß42 stimulated microglia to secrete TNF-a, but with the addition of 1,25-(OH)2D3, TNF-a levels dropped by nearly eight-fold and to near zero values in the presence of both 1,25-(OH)2D3 and SOCS1 and SOCS 3 mimetics. The reduction of the inflammatory …

The Effect Of Gram-Positive Staphylococcus Aureus Cell Wall Components Lipoteichoic Acid And Peptidoglycan On Cytokine Production, Cytoskeletal Arrangement, And Cell Viability On Raw 264.7 Murine Macrophages, Gabrielle String

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In this study, gram positive Staphylococcus aureus cell wall components such as lipoteichoic acid (LTA) and peptidoglycan (PGN) were used to study the potential inflammatory and anti-inflammatory cytokine response, cytoskeletal arrangement and cell viability on RAW264.7 murine macrophages over 24 hours. The effect of S.aureus LTA and PGN (5 [mu]g/mL) on RAW 264.7 macrophages was evaluated every six hours for twenty-four hours. Inflammatory cytokine (TNF-a) production peaked at 6 hours before decreasing over time. Anti-inflammatory cytokine (IL-10) production peaked between 12 and 18 hours. During the first twenty-four hours, cytotoxicity of treated macrophages, as defined as the release of lactate …

The Impact Of Socs1 And Socs3 Peptide Mimetics On Rho And Cdc42 Proteins Expression, F-Actin Cytoskeleton Rearrangements, And Cytokines Production Of Uninfected And Hsv-1 Infected M1 And M2 Raw 264.7 Murine Macrophages, Maha A. Elwardany

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The immune system plays an essential role in the pathogenesis of HSV-1 during the lytic phase of the disease, orchestrating the inflammatory response, retaining the virus in its latent phase and preventing the recurrence of HSV-1 infection. Macrophages display a vital role in the innate and adaptive immune responses during multiple phases of HSV-1 infection. Polarized macrophages are categorized into two distinct classes with diverse functions. The classically activated M1 can engulf and destroy the microbial agents, produce proinflammatory cytokines, and participate in the pathogenesis of many inflammatory diseases. The alternatively activated M2 induces anti-inflammatory mediators and stimulates tissue remodeling …

Effect Of Exposure Of Raw264.7 Macrophages To Salmonella Typhimurium Components On Cell Viability, Cytoskeleton Re-Arrangement And Cytokine Secretion, Khalid Abdullah Alyahya

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Innate immune system plays an important role in individual's protection against pathogens and in activation of adaptive immune system. Utilizing RAW 264.7 murine macrophages as an innate immune response representative in this study, we analyzed the effect of invasive pathogen's components (e.g. flagellin) on the arrangement of macrophage's cytoskeleton, on viability of immune cells and on secretion of pro-inflammatory and anti-inflammatory cytokines and on fluorescence intensity of cytoskeleton after rearrangement. Additionally, we studied the similarity and differences between bacterial (Salmonella typhimurium) and synthetic TLR4 agonist (synthetic lipid-A) on viability, fluorescence intensity, cytokine secretion, and cytoskeleton rearrangements. Similarly, we studied the …

A Novel Molecular Relationship Between Parn And Pld That, When Deregulated, Contributes To The Aggressive Phenotype Of Breast Cancer Cell Lines., Taylor Elaine Miller

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The removal of mRNA transcript poly(A) tails by 3'-5' exonucleases is the rate-limiting step for controlled mRNA decay in eukaryotes. Poly(A)-specific ribonuclease (PARN) is one such exonuclease that degrades poly(A) tails, and although its in vitro activity is well-characterized, PARN’s patho-physiological roles in the cell are not well understood. Prior studies have found a possible role for PARN in cancer, in that PARN expression levels in human breast cancer tissues are often decreased compared to normal control tissues. Indeed, data mined from the ONCOMINE cancer array database showed that PARN is downregulated in patient invasive breast carcinoma samples compared to …

Antimicrobial Activity Of Fractionated Borohydride-Capped And Electrochemical Colloidal Silver, Marjorie M. Markopoulos

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Silver nanoparticles (AgNPs) and ionic silver (Ag+) are known to be broad-spectrum antimicrobial agents. Recent studies show these agents may be an alternative to the most widely used drinking water disinfectant, chlorine. Chlorine is a toxic industrial chemical with a lethal concentration of 430 ppm after 30 minutes. Additionally, chlorine can react with naturally occurring materials to produce a number of disinfection byproducts such as chloroform and trihalomethanes. Some of these byproducts pose cancer risks in addition to other negative impacts to human health. These would be eliminated with the use of Ag+ or AgNPs. The main goal of this …

The Neonatal Anti-Viral Response Fails To Control Measles Virus Spread In Neurons Despite Interferon-Gamma Expression And A Th1-Like Cytokine Profile, Priya Ganesan

Electronic Theses and Dissertations

Neonates are highly susceptible to infections in the central nervous system (CNS) and have a greater risk of viral infections and encephalopathies. Neurotropic viral infections can lead to blindness, hearing loss and neurological deficiencies such as cognitive impairment, epilepsy, and even death in the neonatal and pediatric populations. Viral infections also are hypothesized to indirectly contribute to neurodegenerative and neuropsychiatric diseases such as Schizophrenia and Parkinson’s disease later in life due to early neuronal damage or stress. Many diverse viruses are capable of invading the neonatal CNS including Borna Disease Virus, Coxsackievirus (CV), Herpes simplex viruses (HSV), and measles virus. …

Contribution Of A Class Ii Ribonucleotide Reductase To The Manganese Dependence Of Streptococcus Sanguinis, John L. Smith

Theses and Dissertations

Manganese-deficient Streptococcus sanguinis mutants exhibit a dramatic decrease in virulence for infective endocarditis and in aerobic growth in manganese-limited media. Loss of activity of a manganese-dependent, oxygen-dependent ribonucleotide reductase (RNR) could explain the decrease in virulence. When the genes encoding this RNR are deleted, there is no growth of the mutant in aerobic broth culture or in an animal model. Testing the contribution of the aerobic RNR to the phenotype of a manganese transporter mutant, a heterologous class II RNR from Lactobacillus leichmannii called NrdJ that requires B₁₂ rather than manganese as a cofactor was previously introduced into an …

Hub Proteins, Paralogs, And Unknown Proteins In Bacterial Interaction Networks, Neha Sakhawalkar

Theses and Dissertations

Proteins are the functional units of cells. However, a major portion of the proteome does not have a known functional annotation. This dissertation explores protein -protein interactions, involving these uncharacterized or unknown function proteins. Initially, protein – protein interactions were tested and analyzed for paralogous proteins in Escherichia coli. To expand this concept further and to get an overview, protein – protein interactions were analyzed using ‘comparative interactomics’ for four pathogenic bacterial species including Escherichia coli, Yersinia pestis, Vibrio cholerae and Staphylococcus aureus. This approach was used to study unknown function protein pairs as well as to …

Come Fly With Me: Using Amixicile To Target Periodontal Pathogens And Elucidating The Innate Immune Response In Drosophila Melanogaster, Kathryn Sinclair

Theses and Dissertations

Periodontal diseases (PD) affect 46% of American adults over age 30. These diseases cause symptoms including bleeding and swelling of the gums, bone resorption, and tooth loss, that affect quality of life and have a high economic burden. Periodontal diseases are caused by an imbalance in the oral microbiome, from a healthy state that contains anti-inflammatory commensals like Streptococcus gordonii and mitis, to a diseased state that has pro-inflammatory anaerobic pathogens including Porphyromonas gingivalis, Prevotella intermedia, Fusobacterium nucleatum, and Tannerella forsythia. The latter initiate disease progression in the oral cavity. However, it’s the host immune …

Readiness For Antimicrobial Resistance (Amr) Surveillance In Pakistan; A Model For Laboratory Strengthening, Dania Khalid Saeed, Rumina Hasan, Mahwish Naim, Afia Zafar, Erum Khan, Kauser Jabeen, Seema Irfan, Imran Ahmed Haq, Mohammad Zeeshan, Zabin Wajidali, Joveria Farooqi, Sadia Shakoor, Abdul Chagla, Jason Roy

Department of Pathology and Laboratory Medicine

Background: Limited capacity of laboratories for antimicrobial susceptibility testing (AST) presents a critical diagnostic bottleneck in resource limited countries. This paper aims to identify such gaps and to explore whether laboratory networks could contribute towards improving AST in low resource settings.
Methods: A self-assessment tool to assess antimicrobial susceptibility testing capacity was administered as a pre-workshop activity to participants from 30 microbiology laboratories in 3 cities in Pakistan. Data from public and private laboratories was analyzed and capacity of each scored in percentage terms. Laboratories from Karachi were invited to join a support network. A cohort of five laboratories that …

Frequency Of Colistin And Fosfomycin Resistance In Carbapenem-Resistant Enterobacteriaceae From A Tertiary Care Hospital In Karachi, Salima Qamar, Najma Shaheen, Sadia Shakoor, Joveria Farooqi, Kauser Jabeen, Rumina Hasan

Department of Pathology and Laboratory Medicine

Introduction:
Management of infections with carbapenem-resistant Enterobacteriaceae (CRE) is challenging. In recent times, agents such as colistin and fosfomycin have been used in combination with other antibiotics to treat such infections. In this study, we aim to seek frequency of colistin and fosfomycin resistance in CRE from Pakistan.
Methods:
This study was conducted at clinical laboratories, Aga Khan University Hospital. In total, 251 CRE were included in the study. Colistin minimum inhibitory concentrations (MICs) were performed using broth microdilution (BMD) method and VITEK 2 system, whereas fosfomycin susceptibility was performed using Kirby-Bauer method. MIC50 and MIC90 were calculated for colistin …

Phenotypic Low-Level Isoniazid Resistance As A Marker To Predict Ethionamide Resistance In Mycobacterium Tuberculosis, Salima Qamar, Joveria Q. Farooqi, Kausar Jabeen, Rumina Hasan

Department of Pathology and Laboratory Medicine

Background: Tuberculosis is one of the most prevalent diseases in Pakistan. Pakistan has the highest burden of MDR-TB in the Eastern Mediterranean region. Ethionamide is an anti-tuberculous drug frequently used to treat MDR-TB. Its drug susceptibility testing is not easily available in resource limited settings. Since it acts on the same target protein as isoniazid (inhA protein encoded by inhA gene), we sought to find out if phenotypic isoniazid resistance can be a marker of ethionamide resistance.
Materials and Methods: This was a retrospective observational study conducted at the Aga Khan University hospital section of microbiology. Data was retrieved between …

Utility Of Peripheral Film Findings And Its Correlation With Automated Analyzer - An Audit From Tertiary Care Hospital, Sidra Asad, Imran Ahmed, Natasha Ali

Department of Pathology and Laboratory Medicine

Background and Objective: With the advent of automated hematology analyzer, the use of traditional microscopy of blood film has become limited. The objective of our study was to determine the percentage of peripheral blood smear review in our institution in the era of automation and to identify reasons of manual review.
Materials and Methods: This was a prospective audit from January 1, 2015, to January 15, 2015. Consecutive complete blood count (CBC) samples and peripheral smear requests made up the sample size. All age groups and genders were included. CBCs were performed on Sysmex XE-5000. The variables to be analyzed …

Papillary Craniopharyngioma: A Clinicopathologic Study Of A Rare Entity From A Major Tertiary Care Center In Pakistan, Muhammad Usman Tariq, Nasir Ud Din, Zubair Ahmad, Wasim Memon

Department of Pathology and Laboratory Medicine

Background: Papillary craniopharyngioma (PCP) are uncommon variants of craniopharyngiomas (CP), which are benign epithelial neoplasms of the sellar and suprasellar region. Histologically, PCPs are typically composed of well-differentiated stratified squamous epithelium; however, focal variations are not uncommon. A distinction from other lesions of the region, despite being difficult to achieve due to the overlapping radiological and clinical features, is important for adequate treatment to be administered.
Objective: Our aim was to study the clinical and histological features of PCP with emphasis on features that are helpful in its distinction from other lesions that are similar in appearance.
Materials and Methods: …

Rapid Detection Of In Vitro Antituberculous Drug Resistance Among Smear-Positive Respiratory Samples Using Microcolony Detection-Based Direct Drug Susceptibility Testing Method, Irim Iftikhar, Seema Irfan, Joveria Farooqi, Zahida Azizullah, Rumina Hasan

Department of Pathology and Laboratory Medicine

Background: With the rise in multidrug-resistant tuberculosis, there is a search for newer techniques that will rapidly detect drug-resistant Mycobacterium tuberculosis. Although molecular techniques can detect resistance, culture is still considered gold standard, especially in resource-limited settings where quick, cheap, and easy techniques are needed. The aim of the study was to evaluate microcolony method thin layer agar (TLA) for quick detection of resistance against the first- and second-line antituberculous drugs in clinical isolates. This was a cross-sectional study performed at Aga Khan University Hospital.
Material and Methods: A total of 87 Z-N stain smear-positive pulmonary samples were received and …

Studies Relating Pqq Biosynthesis To Putative Peptidases And Operon Structure In Pseudomonas Species, Benjamin Diaz

Theses and Dissertations--Plant and Soil Sciences

Several bacteria isolated from the broccoli rhizosphere were assayed to compare their ability to solubilize phosphate and release pyrroloquinoline quinone (PQQ) into the surrounding media. Subsequently, their genomes were sequenced and analyzed for PQQ biosynthesis operon structure. PQQ biosynthesis genes pqqA-F were found in all isolates. The order of PQQ biosynthesis genes and predicted amino acid sequences were compared to each other and the host’s ability to solubilize phosphate and release PQQ. In all Pseudomonas species, two putative protease genes, pqqF, and pqqG, flanked the canonical pqqA-pqqE biosynthesis operon. No mechanistic studies have confirmed the function of …

A Systems Biology Approach For Predicting Essential Genes And Deciphering Their Dynamics Under Stress In Streptococcus Sanguinis, Fadi El-Rami

Theses and Dissertations

Infectious diseases are the top leading cause of death worldwide. Identifying essential genes, genes indispensable for survival, has been proven indispensable in defining new therapeutic targets against pathogens, major elements of the minimal set genome to be harnessed in synthetic biology, and determinants of evolutionary relationships of phylogenetically distant species. Thus, essentiality studies promise valuable revenues that can decipher much of biological complexities.

Taking advantage of the available microbial sequences and the essentiality studies conducted in various microbial models, we proposed a framework for the prediction of essential genes based on our experimentally verified knowledge of the pathways involved in …