Life Sciences Commons^™

Functional Analysis Of Legionella Pneumophila Effector Protein, Shreya Neupane

Masters Theses & Specialist Projects

Legionella pneumophila is a gram-negative bacterium that causes Legionnaire’s disease (a severe form of pneumonia) in humans. L. pneumophila can cause infection by utilizing its Type IV secretion system, a protein secretion system that transports proteins from the bacterial cytosol into the infected macrophage. Effectors released from the Type Iv secretion system allow L. pneumophila to create a safe environment to survive, replicate and cause infection. One such effector, RavQ, inhibits cell proliferation of mammalian HEK 293T cells and localizes to the cell’s nucleus, leading us to hypothesize that RavQ interferes with cellular activity in the nucleus. To detect its …

Functional Characterization Of Mycobacterium Smegmatis Phage Moomoo Gene Products: Identification Of Toxic Genes, Whitney Heard

Masters Theses & Specialist Projects

Mycobacteriophages are a group of phages that infect members of the genus Mycobacteria. Previous studies have conducted extensive comparisons of the mycobacteriophage genomes and amino acid sequences to establish different phage families. MooMoo is a singleton mycobacteriophage that has been characterized due to its lack of appreciable homology to other phages. Some of its unique properties include its structure and the isolation of a mutant that causes clear plaque phenotypes. The experiments described in this thesis identified three MooMoo phage (gp87, gp90, and gp91) encoded proteins that are toxic to the bacterial host, Mycobacterium smegmatis.Through the use of a …

Improved Antimicrobial Properties Of Silver Nanoparticles With Methylene Blue, Ermek Belekov

Masters Theses & Specialist Projects

Photosensitizing agents are the cornerstone of photodynamic therapy (PDT) that play essential role in deactivation process of multidrug resistant pathogens and tumor treatments. In this work we studied a photosensitizing agent made from mixture of silver nanoparticles (Ag NPs) and methylene blue (MB) which possess improved important characteristics like high photostability and high singlet oxygen yield. Ag NPs were synthesized by pulsed laser ablation technique in different aqueous solutions like Polyvinylpyrrolidone (PVP), citrate and Deionized (DI) water. The synthesized Ag NPs were characterized in depth using with transmission electron microscopy (TEM), UVVisible (UV-Vis), and photoluminescence (PL) spectra. Ag NPs were …

Properties Of A Genetically Unique Mycobacteriophage, Amanda K. Staples

Masters Theses & Specialist Projects

Bacteriophage MooMoo is a temperate phage that was isolated and propagated on Mycobacterium smegmatis (M. smeg). It typically produces turbid plaques, however spontaneous clear plaque mutants can be readily isolated. Both turbid (MooMoo-T) and clear plaque (MooMoo-C) formers can establish stable lysogens, but the parental turbid plaque forming phage has a higher lysogenic frequency. The phage repressor protein typically plays the central role in regulating the lysis/lysogeny decision. Therefore, we expected that the mutation responsible for the clear plaque phenotype would be located in the repressor gene. Remarkably, whole genome sequencing detected a single base pair mutation in the minor …

Synthesis And Kinetic Studies Of High-Valent Metal-Oxo Species Generated By Photochemical And Chemical Methods, Haiyan Liu

Masters Theses & Specialist Projects

Highly reactive iron-oxo intermediates play important roles as active oxidants in enzymatic and synthetic catalytic oxidation. Many transition metal catalysts are designed for biomimetic studies of the predominant oxidation catalysts in Nature, namely cytochrome P450 enzymes. In this work, a series of iron(IV)-oxo porphyrins [Fe^IV(Por)O] and manganese(IV)-oxo porphyrins [Mn^IV(Por)O] have been successfully produced in two electron-deficient ligands by photochemical and chemical methods, and spectroscopically characterized by UV-vis, and ¹H-NMR.

With iodobenzene diacetate [PhI(OAc)₂] as the oxygen source, iron(III) porphyrin and manganese(III) porphyrin complexes converted to the corresponding metal(IV)-oxo species as oxygen atom transfer …

Sequence Extension Of The Tryptophan And Shikimate Operons In Clostridium Scatologenes Atcc 25775, Shawn Johnston Smiley

Masters Theses & Specialist Projects

3-Methylindole and 4-methylphenol are cytotoxic and malodorant compounds derived from tryptophan and tyrosine, respectively. Each is present in swine waste lagoons and contributes to malodorous emissions from agricultural facilities. Clostridium scatologenes ATCC 25775 produces both compounds and serves as a model organism to study their metabolism and function. Through the repeated assembly and annotation of the Clostridium scatologenes genome, we propose a novel pathway for tryptophan degradation and 3-methylindole production by this organism. The genome of Clostridium scatologenes was sequenced, and re-assembled into contigs. Key elements of the tryptophan and shikimate pathways were identified. Contigs containing these elements were extracted …

Detection And Quantitation Of Tetracycline Antibiotics In Agricultural Swine Wastes, Ali Jamal Abdulrheem

Masters Theses & Specialist Projects

The spread of tetracyclines through agricultural systems is causing the present bacteria to develop antibiotic resistance. The spread of this bacteria, as well as the tetracycline antibiotics in the environment is dangerous because these antibiotics pose health hazards for humans. The overuse of antibiotics, which are added to livestock feed, results in the antibiotics being released into the environment via animal feces. In this research, we have attempted to design an analytical method to isolate antibiotics from agricultural wastes with subsequent detection using liquid chromatography and mass spectrometry (LC-MS). The antibiotics investigated in this study were tetracycline, chlortetracycline, and oxytetracycline. …

Farm-To-Fork Fresh Produce Food Safety: An Evaluation Of Perceptions, Knowledge, And Implementation Of Good Agriculture Practices In Kentucky, Daniel J. Sinkel

Masters Theses & Specialist Projects

Farmers’ markets have increasingly become a popular venue for purchase of fresh, locally-grown produce, with the number of farmers’ markets in Kentucky reaching an all-time high of 159 in 2016. Good Agriculture Practices (GAPs) is a program created by the USDA’s Agriculture Marketing Service to function as a food safety audit for small-scale fresh produce growers, such as those who sell fresh produce at local farmers’ markets. However, under the provisions of the Food Safety Modernization Act of 2011, small-scale farmers who sell an average of $25,000 in annual fresh produce sales across the span of three years are exempt …

Decay Of Macroalgae And Leaves And Their Relation To Detrital Food Webs, Megan E. Grandinetti

Masters Theses & Specialist Projects

This project addressed if decaying macroalgae and leaf detritus play a major role in the detrital pool of a 7th-order karst riverine system. Decay rates, macroinvertebrates colonization patterns, and change in δ13C values of Cladophora, Platanus occidentalis, and a mix of Acer negundo and A. saccharinum were tracked during summer and autumn months for portions of multiple years.

Packs of air-dried Cladophora, Acer, and P. occidentalis were placed in mesh bags and put in groups (n=4) in wire baskets. Seven baskets were submerged in riffle (0.5 m) and deeper run (2 m) habitats. Benthic organic matter was collected with each …

Epiphytic Diatom Community Structure In A Karst Riverine System, Gregory John Barren

Masters Theses & Specialist Projects

The goal of this study was to assess the epiphytic diatom community structure of two host species along a karst gradient in the upper Green River, Kentucky to a gain a better understanding of the role of diatoms in the food web. The host species studied were Podostemum ceratophyllum and Cladophora. Percent cover of P. ceratophyllum and Cladophora were quantified in the four study reaches. The host species were sampled near-shore and mid-channel in each reach in September and October of 2013. After diatoms were extracted from the host and enumerated the density and diversity were quantified. Twelve genera were …

Defining The Requirements For Early Gene Expression In Bacteriophage Hk639, Amanda L. Seaton

Masters Theses & Specialist Projects

Lambdoid phages suppress transcription termination to fully express their genes. Antitermination of early gene expression in most lambdoid phages is mediated by an interaction between the N protein and a number of host-encoded factors. Bacteriophage HK022 does not rely on a protein for antitermination. To promote full expression of early phage genes, the transcripts of the HK022 put sites interact directly with RNA polymerase to convert it to a termination resistant form. Bacteriophage HK639 also uses RNA-mediated antitermination. However, it only possesses a single put-like element in its left operon. Because most lambdoid phages, including HK022, have antiterminator elements in …

Single Step Synthesis Of Antibiotic Kanamycin Embedded Gold Nanoparticles For Efficient Antibacterial Activity, Shravan Gavva

Masters Theses & Specialist Projects

Nanotechnology has become the most advanced type of drug delivery system within the last decade. This advancement shifted the focus on small carriers to increase the efficiency of the drugs. Among these, gold nanoparticles (GNPs) were found to have profound biomedical applications. In current research, kanamycin embedded GNPs were prepared in a single step, single phase, and bio-friendly (green synthesis) procedure. The synthesized Kanamycin-GNPs (Kan-GNPs) were spherical in shape and had a size range of 15 ± 3 nm. The chosen kanamycin is an aminoglycosidic antibiotic that is isolated from Streptomyces kanamyceticus. These special antibiotic GNPs are further characterized using …

Production And Degradation Of 4-Ethylphenol In Lactobacillus Sp. Pep8 Cultures And In Blended Swine Lagoon Enrichments, Clinton W. Copp

Masters Theses & Specialist Projects

4-Ethylphenol (4-EP) is a malodorant of swine waste and is derived from a component of lignin called p-coumaric acid (p-CA). The production of 4-EP from lignin in swine waste is untested. Additionally, the effect of Fe (III) on 4-EP levels is unknown. Four experiments were performed to determine if Lactobacillus sp. pep8 cultures, as well as enriched swine lagoon slurries, could liberate p-CA from lignin and convert p-CA to 4-EP. Furthermore, it was tested if the addition of Fe (III) influences the conversion of p- CA to 4-EP.

Experiment 1 tested Lactobacillus sp. pep8 cultures to determine if the addition …

Two-Phase Partitioning System Using Elvax 40w Polymer For The Biodegradation Of Aqueous Phenols, Amit Suresh Ghode

Masters Theses & Specialist Projects

A solid-liquid two phase partitioning system (TPPS) is a new technology platform for destroying toxic organic compounds. TPPS have traditionally been operated by using an immiscible organic phase which partitions organic compounds into the aqueous phase. TPPS using an immiscible organic phase suffers from several limitations such as the organic phase could be biodegradable and hence only certain compatible microbial strains could be used. This therefore, eliminates the desired use of mixed microbial populations for efficient degradation. A solid-liquid two phase partitioning system, in which solid polymeric beads replace liquid organic phase, appears to have benefits over the traditional liquid-liquid …

Analysis Of The Clear Plaque Phenotype Of The Bacteriophage Hk75, Phani Chandrika Kunapuli

Masters Theses & Specialist Projects

The growth of bacteriophage HK75 is inhibited by specific mutations in the zinc binding domain of the host RNA polymerase beta prime subunit. It shares this rare property with bacteriophage HK022 and other phages that use RNA mediated antitermination to promote early gene expression. Recent genomic analysis of HK75 and HK022 has confirmed the relatedness of these two phages and place HK75 in the lambdoid family of bacteriophages. Lambdoid phages are temperate and can adopt a lytic or lysogenic lifestyle upon infection of a suitable host. However, HK75 only forms clear plaques and thus appears to be defective in its …

Adding Upstream Sequence And A Downstream Reporter To The Bile Acid Inducible Promoter Of Clostridium Scindens Vpi 12708, Bryan Patrick Mason

Masters Theses & Specialist Projects

Bile acids in the small intestines of animals serve to breakdown fats and fatsoluble vitamins. Most of the bile acids are reabsorbed into the enterohepatic circulation, but approximately five percent of these bile acids pass into the large intestine. These bile acids are swiftly deconjugated by the bacterial population, and then subjected to further intestinal bacterial chemical modifications. The most significant of these modifications are 7α-dehydroxylations which form secondary bile acids (deoxycholate and lithocholate). Much research has illuminated the 7α-dehydroxylation pathway: of particular interest is the bile acid inducible operon, for which Clostridium scindens VPI 12708 serves as the model …

Estimation Of Microbial Diversity In Poultry Litter Using Terminal Restriction Fragment Length Polymorphism And Isolation Of Phosphate Accumulating Bacteria From Poultry Litter, Yoganand Vadari

Masters Theses & Specialist Projects

The contamination of fresh water by phosphates in poultry litter results in substantial eutrophication of fresh water causing fish kills and other types of environmental damage. The poultry indus try in Kentucky is expanding rapidly. The number of broilers is increasing as more poultry farms are established in the state producing waste that needs disposal. Investigations were made to study the possibility of using microorganisms normally found in poultry litter to sequester phosphate, thereby delaying phosphate runoff after litter is applied to croplands. Little is known, however, about the microflora of poultry litter. Terminal restriction fragment length polymorphism of 16S …

A Modified Scheme For The Isolation And Enumeration Of Bacteria In Municipal Sewage Sludge, Kelly Ball

Masters Theses & Specialist Projects

Because of the potential health hazards associated with the use of sludge for agricultural purposes, Dudley et al (1980) published a scheme for the routine analysis of bacteria in municipal sewage sludge. In this study, the Dudley et al scheme (1980) was modified by updating some of the procedures. Aerobically digested sludge generated by the Bowling Green Wastewater Treatment Plant, Bowling Green, Kentucky, was analyzed using the modified scheme. Sludge samples were collected once every two months over a one-year period from October 1989 to August 1990.

Egg yolk-free tryptose sulfite cycloserine agar in conjunction with the revewrse CAMP test …

A Preliminary Investigation Of The Effect Of Storm Events On The Concentration Of Volatile Organic Compounds In The Lost River Cave System, Warren County, Kentucky, Francis Michael Cretella

Masters Theses & Specialist Projects

Samples of cave water were analyzed for volatile organic compounds during four separate storm events. The major compounds detected were toluene, 1,1,1-trichloroethane, trans-1,2-dichloroethylene, methylene chloride, and 1,1-dicloroethane. Minor compounds detected were trichloroethylene, tetrachloroethylene, 1,1-dichloroethylene, and ethylbenzene. These compounds were detected in varying amounts at each of the three study sites. During a storm event, the levels of each contaminant changed significantly at all three sites studied. Two effects were observed when a storm event occurred. The first effect was the dilution of the volatile organic compounds during the storm event. These effects were related to the amount of rainfall and …

A Bacteriological And Chemical Analysis Of Nonpoint Source Pollution In A Karst Aquifer Bowling Green, Kentucky, Wayne Green

Masters Theses & Specialist Projects

Monthly water samples collected from four sites in the Lost River Groundwater Basin, a shallow karst aquifer in the Bowling Green-Warren County area of Kentucky, represented samples from sites receiving conduit and diffuse flow. All sites were severely contaminated with bacteria, and on some occasions the surface water criteria for some heavy metals were exceeded.

Of the total 334 bacterial colonies identified 92.1% were verified as Escherichia coli by the API20E system. Acinetobacter calcoaceticus var. anitratum accounted for 2.10% of colonies; Citrobacter freundii for 0.30% Klebsiella pneumoniae for 0.90%; Klebsiella oxytoca, 0.90%; Citrobacter amalonaticus 0.30%; Enterobacter cloacae, 1.20%; …

Description And Seasonal Variation In Incidence Of A New Species Of Myxosporidian Parasite (Class Myxosporidea) Of The Bluegill Sunfish, Lepomis Macrochirus Rafineque, In Kentucky, Stephen Bayes Crider

Masters Theses & Specialist Projects

A new histozoic myxosporidian parasite (Class Mysoxporidea), Myxobolus meglitschi sp. nov., infecting young-of-the-year and yearling bluegill sunfish, Lepomis macrochirus Raf., from Shanty Hollow Lake, Warren County, Kentucky was described. Phenology was investigated from January 20 to December, 1969.

The magnitude of infection varied seasonally in bluegill. Incidence was highest in July and August (43.92) in host populations, declining, but not disappearing from September to December 20, 1969. The infection exhibited a yearly mean incidence of 22.318.

The pattern of distribution of cysts on hosts varied seasonally. During periods of low incidence cysts were confined primarily to a postanal area below …

Investigation In The Physiology Of The Spore Formation Of Pseudoples Trifolii (Rostr.) And Petr., Emory S. Crosby

Masters Theses & Specialist Projects

Plant pathologists have worked to some extent for many years in an attempt to describe the complete physiology of the organism responsible for pepper spot of clover, particularly as found on white clover (Trifolium repens, L.). These works have not been extensive or concentrated and at the present time, no definite conclusions have been reached.

The causal agent of pepper spot of clover is Pseudoples trifolii (Rostr.).

Pseudoples trifolii usually is found parasitizing clover in the early spring and continues throughout the spring until the temperature exceeds thirty degrees centigrade. Elliott established that Pseudoples trifolii will not produce infectious spores …