View All Items

Title: BACTERIA THAT RESIST CENTRIFUGAL FORCE.
Creator: Kessler, Nickolas, Moore, Sean, University of Central Florida
Abstract / Description: Our lab discovered that approximately 1 in 10,000 Escherichia coli cells in stationary phase remain in suspension after a high g-force centrifuge event. To establish the mechanism behind this curious phenotype, multiple mutant strains of E. coli were independently evolved such that the majority of their populations resisted migration when exposed to high centrifugal forces. Genomic DNA sequencing of the mutants' revealed unique, isolated mutations in genes involved in capsule synthesis and...
Show moreOur lab discovered that approximately 1 in 10,000 Escherichia coli cells in stationary phase remain in suspension after a high g-force centrifuge event. To establish the mechanism behind this curious phenotype, multiple mutant strains of E. coli were independently evolved such that the majority of their populations resisted migration when exposed to high centrifugal forces. Genomic DNA sequencing of the mutants' revealed unique, isolated mutations in genes involved in capsule synthesis and exopolysaccharide (EPS) production. Each mutant exhibits a novel mechanism that allows them to remain in suspension. The mutants were further characterized by determining their growth rates, strengths of resistance to various centrifugal forces, the phenotype's dependence on a carbon source, and timing of the phenotype's presentation. The results revealed: comparable mutant generation times to the wild-type strain, variable resistance to centrifugal force, phenotype dependence on carbon source, and phenotype presentation during early stationary phase. To interrogate the mechanism by which these cells stay in suspension the production of EPS was quantified, and gene knock-outs were performed. Quantification of the EPS revealed approximately a seventeen-fold increase in EPS in the mutants' compared to the wild-type strain. Gene knock-outs revealed the EPS produced can be attached to the outer-membrane or freely secreted into the media by different mechanisms. In addition, this mechanism was further confirmed to be responsible for the centrifuge resistant trait by attaching extracted EPS to polystyrene microspheres. Experimental results show that mutant extracted EPS treated beads caused increased bead retention in suspension compared to wild-type EPS treated beads. These results reveal that E. coli is using a novel mechanism to adapt to a new environmental factor introduced to remove the bacteria. With the discovery of this mechanism and the transferability to inorganic objects industrial applications are now envisioned where particle sedimentation is controllable and mixtures remain homogenized by attaching optically transparent biomolecules.
Show less
Date Issued: 2018
Identifier: CFH2000332, ucf:45800
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH2000332

Title: THE EFFECT OF MISMATCH PRIMERS ON THE EFFICIENCY OF AMPLIFICATION IN QUANTITATIVE POLYMERASE CHAIN REACTIONS.
Creator: Dawkins, Molly C, Moore, Sean, University of Central Florida
Abstract / Description: Polymerase chain reaction (PCR) is a method used in many research protocols to amplify a small amount of a short segment of DNA to millions of copies. PCR is used for many taxonomic studies, as well as for some medical diagnostic procedures. Through PCR, short DNA primers bind to the template DNA to allow the thermostable DNA polymerase to copy the DNA. Often, researchers create universal primers to target a conserved region of DNA in multiple species, for example, the 16S rRNA gene in...
Show morePolymerase chain reaction (PCR) is a method used in many research protocols to amplify a small amount of a short segment of DNA to millions of copies. PCR is used for many taxonomic studies, as well as for some medical diagnostic procedures. Through PCR, short DNA primers bind to the template DNA to allow the thermostable DNA polymerase to copy the DNA. Often, researchers create universal primers to target a conserved region of DNA in multiple species, for example, the 16S rRNA gene in bacteria. The problem with these universal primers is that they do not always perfectly match the target DNA. The mismatch primers can still bind to the template, but could affect the efficiency of the PCR amplification. The effect of mismatch primers on the efficiency of the amplification in PCR is the focus of this thesis. Four forward primers with various mismatch overhangs were generated and incorporated into a DNA template through an initial PCR. These primers contained the binding region complementary to the V3/V4 region of the 16S rRNA bacterial gene. Further quantitative PCR (qPCR) reactions were run on these newly-made templates using two sets of primers complementary to the 16S rRNA gene region – one with ambiguous base pairs, one with unambiguous base pairs. The qPCR amplification curves, the Cq values, and the initial concentrations of DNA products (seed values) were analyzed and compared. The results showed differences in the Cq values and seed values between the reactions containing mismatches and those not containing mismatches. Other variables including annealing temperature, addition of Illumina sequencing tails to the primers, and initial primer concentration were also tested to determine if these variables had an effect on the amplification. The results from these reactions using different variables were inconclusive.
Show less
Date Issued: 2018
Identifier: CFH2000361, ucf:45761
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH2000361

Title: EXPRESSION LEVELS OF VIRULENCE GENES IN GROUP A STREPTOCOCCI: A RESPONSE TO AEROSOLIZED PROPYLENE GLYCOL.
Creator: Costello, Michael S, Hoffman, Eric, Moore, Sean D., University of Central Florida
Abstract / Description: Electronic cigarette usage is becoming increasingly prevalent among school age children and young adults. A known bactericidal agent, propylene glycol, is often used as a carrier for nicotine, flavoring, and additional constituents of electronic cigarette juice. This study examined the relationship between propylene glycol and virulence gene expression in Streptococcus pyogenes, a respiratory tract pathogen commonly found in school-age individuals. A variety of virulence genes controlled by...
Show moreElectronic cigarette usage is becoming increasingly prevalent among school age children and young adults. A known bactericidal agent, propylene glycol, is often used as a carrier for nicotine, flavoring, and additional constituents of electronic cigarette juice. This study examined the relationship between propylene glycol and virulence gene expression in Streptococcus pyogenes, a respiratory tract pathogen commonly found in school-age individuals. A variety of virulence genes controlled by the three stand alone regulators mga, RofA, and Rgg/RopB were sampled in an effort to understand the pathway by which virulence is affected. The genes chosen encode C5a peptidase, fibronectin binding protein, hyaluronate lyase, NAD glycohydrolase, Streptococcal pyrogenic exotoxin A and B, streptodornase, streptokinase, Streptolysin O, and Streptolysin S. No significant change in gene expression was observed, but a novel method to test the effects of aerosols on cells was developed. This method can be used in the future to observe the effect of aerosols, including commercial electronic cigarette juice, on both bacterial and mammalian cells.
Show less
Date Issued: 2016
Identifier: CFH2000106, ucf:46048
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH2000106

Title: The microbial ecosystem of beer spoilage and souring: Competition and cooperation in the age of bioinformatics.
Creator: Kettring, Andrew, Moore, Sean, Cole, Alexander, Self, William, University of Central Florida
Abstract / Description: The brewing industry generates $350 billion in revenue in the US annually, representing 1.9% of the gross domestic product. Spoilage is a persistent problem throughout production and distribution that causes economic loss, and is therefore meticulously avoided. Contrarily, artisanal sour beers are necessarily produced by a diverse variety of these spoilage organisms metabolically interacting in symbiosis as a microbial ecosystem. We sought to gain insight into factors driving assembly of...
Show moreThe brewing industry generates $350 billion in revenue in the US annually, representing 1.9% of the gross domestic product. Spoilage is a persistent problem throughout production and distribution that causes economic loss, and is therefore meticulously avoided. Contrarily, artisanal sour beers are necessarily produced by a diverse variety of these spoilage organisms metabolically interacting in symbiosis as a microbial ecosystem. We sought to gain insight into factors driving assembly of microbial communities by testing a long-debated Darwinian hypothesis. A collection of community members were screened in co-culture and novel bioinformatics tools were developed to predict observed interactions. A fundamental understanding of these relationships is paramount to beer production and sets a precedent for the study of similar microbial communities that impact human health.
Show less
Date Issued: 2017
Identifier: CFE0007288, ucf:52147
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007288

Title: Exacerbation of efp- sickness in Escherichia coli by an uncharacterized RNA helicase.
Creator: Wingo, Robert, Moore, Sean, Roy, Herve, Teter, Kenneth, University of Central Florida
Abstract / Description: In Escherichia coli, growth is rate-limited by translation capacity. Stalled ribosomes have profound effects on a cell such as altered mRNA abundance, decreased ribosome availability, and an imbalanced proteome. The absence of elongation factor P (EF-P), a universally conserved transpeptidation enhancer, presents an extreme example of this scenario, wherein ribosomes accumulate disproportionately onto messages that are more slowly translated and cell growth becomes notably impaired. We...
Show moreIn Escherichia coli, growth is rate-limited by translation capacity. Stalled ribosomes have profound effects on a cell such as altered mRNA abundance, decreased ribosome availability, and an imbalanced proteome. The absence of elongation factor P (EF-P), a universally conserved transpeptidation enhancer, presents an extreme example of this scenario, wherein ribosomes accumulate disproportionately onto messages that are more slowly translated and cell growth becomes notably impaired. We discovered that faster-growing cells arise spontaneously in ?efp cultures, suggesting that translation defects could be circumvented by mutating other genes. This thesis presents a genetic and biochemical analysis of a mechanism ?efp cells employ to overcome translation stress. Using a dual luciferase reporter system, we found that transpeptidation remained hindered in the faster growing ?efp cells. Whole genome sequencing of several fast-growing strains revealed mutations in a poorly characterized RNA helicase called HrpA. We determined that deletion of hrpA, or mutations at several conserved residues critical for HrpA's function, was sufficient to improve the fitness of ?efp cells. HrpA is a DEAH-box RNA helicase and represents a large class of enigmatic proteins that use ATP to restructure cellular RNAs; however, it's direct function in cellular physiology has yet to be clearly demonstrated. Several HrpA mutants were engineered to interrogate the molecular mechanism of HrpA and how its function impairs ?efp cells. Complementation in ?efp ?hrpA cells showed that a number of these mutants were unable to restore sickness, suggesting they were defective in key aspects of RNA processing. It was discovered that wild-type HrpA is associated with actively translating ribosomes and several of the inactive HrpA mutants impose substantial deleterious effects on translation and ribosome production. In sum, the work presented here describes a mechanism by which cells overcome translation stress involving a novel genetic and biochemical relationship between EF-P and HrpA.
Show less
Date Issued: 2018
Identifier: CFE0007267, ucf:52178
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007267

Title: Selenium vs. Sulfur: Investigating the Substrate Specificity of a Selenocysteine Lyase.
Creator: Johnstone, Michael, Self, William, Roy, Herve, Moore, Sean, University of Central Florida
Abstract / Description: Selenium is a vital micronutrient in many organisms. While traces are required for survival, excess amounts are toxic; thus, selenium can be regarded as a biological (")double-edged sword("). Selenium is chemically similar to the essential element sulfur, but curiously, evolution has selected the former over the latter for a subset of oxidoreductases. Enzymes involved in sulfur metabolism are less discriminate in terms of preventing selenium incorporation; however, its specific incorporation...
Show moreSelenium is a vital micronutrient in many organisms. While traces are required for survival, excess amounts are toxic; thus, selenium can be regarded as a biological (")double-edged sword("). Selenium is chemically similar to the essential element sulfur, but curiously, evolution has selected the former over the latter for a subset of oxidoreductases. Enzymes involved in sulfur metabolism are less discriminate in terms of preventing selenium incorporation; however, its specific incorporation into selenoproteins reveals a highly discriminate process that is not completely understood. In this work, we add knowledge to the mechanism for selenium-over-sulfur specificity in hopes of further understanding the controlled regulation of selenium trafficking and the prevention of its toxicity. We have identified SclA, a selenocysteine lyase in the nosocomial pathogen, Enterococcus faecalis, and characterized its enzymatic activity and specificity for L-selenocysteine over L-cysteine. Human selenocysteine lyase contains a residue, D146, which plays a significant role in determining its specificity. A D146K mutation eliminated this trait, allowing non-specific L-cysteine degradation. Using computational biology, we identified an orthologous residue in SclA, H100, and generated mutant enzymes with site-directed mutagenesis. The proteins were overexpressed, purified, and characterized for their biochemical properties. All mutants exhibited varying levels of activity towards L-selenocysteine, hinting at a catalytic role for H100. Additionally, L-cysteine acted as a competitive inhibitor towards all enzymes with higher affinity than L-selenocysteine. Finally, our experiments revealed that SclA possessed extremely poor cysteine desulfurase activity with each mutation exhibiting subtle changes in turnover. Our findings offer key insight into the molecular mechanisms behind selenium-over-sulfur specificity and may further elucidate the role of selenocysteine lyases in vivo.
Show less
Date Issued: 2019
Identifier: CFE0007659, ucf:52481
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007659

Title: Allelic characterization and novel functions of the outer membrane porin U in Vibrio cholerae.
Creator: Sakib, Sk Nazmus, Almagro-Moreno, Salvador, Moore, Sean, Roy, Herve, University of Central Florida
Abstract / Description: Vibrio cholerae is the etiological agent of the severe diarrheal disease cholera. The bacterium is a natural inhabitant of brackish and estuarine waters . To date, only a subset of V. cholerae strains, those belonging to the pandemic group (PG), can cause cholera in humans while the rest (environmental group, EG) cannot cause the disease. Recently, we discovered that V. cholerae PG contains allelic variations in core genes that confer preadaptation to virulence, which we termed Virulence...
Show moreVibrio cholerae is the etiological agent of the severe diarrheal disease cholera. The bacterium is a natural inhabitant of brackish and estuarine waters . To date, only a subset of V. cholerae strains, those belonging to the pandemic group (PG), can cause cholera in humans while the rest (environmental group, EG) cannot cause the disease. Recently, we discovered that V. cholerae PG contains allelic variations in core genes that confer preadaptation to virulence, which we termed Virulence Adaptive Polymorphisms (VAPs). We identified nine core genes that encode potential VAPs, one of which encodes the outer membrane porin U (OmpU). OmpU provides tolerance to bile and acidic pH, resistance to antimicrobials and facilitates biofilm formation. In this study, several alleles of ompU were analyzed to determine whether these VAPs encode different functional properties. We performed multiple phenotypic assays and observed increased survival for strains encoding the PG-like alleles in the presence of bile, organic acid, anionic detergents and the antimicrobial peptide P2. On the other hand, EG-like alleles only showed increased biofilm formation. Interestingly, tests for motility and tolerance of inorganic acid, polymyxin B and protamine sulphate showed no differences in survival for strains encoding either alleles indicating that some of the properties conferred by OmpU are allelic independent. We have also discovered that V. cholerae OmpU shows resistance against Rifamycin, EDTA and Trifluoperazine and interestingly, Rifamycin has been found to be PG-allele dependent. Our findings provide further evidence that genetic variations in core genes lead to the emergence of virulence adaptive traits in pathogenic V. cholerae and can be extrapolated to other bacterial pathogens.
Show less
Date Issued: 2019
Identifier: CFE0007720, ucf:52420
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007720

Title: It takes two to tango: the toxin-chaperone relationship.
Creator: Kellner, Alisha, Teter, Kenneth, Moore, Sean, Cole, Alexander, Harper, James, University of Central Florida
Abstract / Description: Cholera toxin (CT) enters the cell via receptor-mediated endocytosis and travels in a retrograde fashion to the endoplasmic reticulum (ER). The catalytic A1 subunit (CTA1) is then displaced from the rest of the holotoxin, unfolds, and is exported to the cytosol where it regains an active conformation for the ADP-ribosylation of its G-protein target. We have shown that the cytosolic chaperones Hsp90 and Hsc70 are required for CTA1 translocation to the cytosol. We have also shown that both are...
Show moreCholera toxin (CT) enters the cell via receptor-mediated endocytosis and travels in a retrograde fashion to the endoplasmic reticulum (ER). The catalytic A1 subunit (CTA1) is then displaced from the rest of the holotoxin, unfolds, and is exported to the cytosol where it regains an active conformation for the ADP-ribosylation of its G-protein target. We have shown that the cytosolic chaperones Hsp90 and Hsc70 are required for CTA1 translocation to the cytosol. We have also shown that both are able to independently bind and refold CTA1. Using libraries of CTA1-derived peptides, we have identified a single Hsc70 binding site, YYIYVI (CTA1 83-88), within the 192 amino acid protein, as well as two distinct Hsp90 binding sites: an N-terminal RPPDEI (CTA111-16) motif and a C-terminal LDIAPA (CTA1 153-158) motif. The LDIAPA motif is unique to CTA1, but an RPPDEI-like motif is present in four other ER-translocating ADP-ribosylating toxins: pertussis toxin, Pseudomonas aeruginosa exotoxin A, Escherichia coli heat-labile toxin, and Salmonella typhimurium ADP-ribosylating toxin. Using site-directed mutagenesis to further investigate the RPPDEI motif, we found that a modification of either proline residue blocks CTA1 translocation to the cytosol. Our work has identified, for the first time, specific amino acid sequences that are recognized by Hsp90/Hsc70 and are essential for toxin translocation from the ER to the cytosol. CT does not require prolyl isomerases for cellular activity, as is the case for Hsp90-dependent endosome-translocating toxins. We therefore hypothesize that the one or both of the prolines within the RPPDEI motif of CTA1 undergo an isomerization event as CTA1 unfolds in the ER. Furthermore, we predict that the trans- to cis- conformational change of proline(s) is the molecular determinate for the atypical Hsp90 interaction observed with CTA1 and related toxins. Additionally, we have identified Hsp90 and other host factors required for the translocation of pertussis toxin.
Show less
Date Issued: 2019
Identifier: CFE0007661, ucf:52500
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007661

Title: Chlamydia trachomatis Transformants Show a Significant Reduction in Rates of Invasion upon Removal of Key Tarp Domains.
Creator: Parrett, Christopher, Jewett, Travis, Roy, Herve, Moore, Sean, University of Central Florida
Abstract / Description: Chlamydia trachomatis is an obligate, intracellular bacterium which is known to cause multiple human infections including nongonococcal urethritis (serovars D-K), lymphogranuloma venereum (serovars L1, L2, L3) and trachoma (serovars A-C). The infectious form of the bacterium, called the elementary body (EB), harbors a type III secreted effector known as Tarp (translocated actin recruiting phosphoprotein) which is a candidate virulence factor and is hypothesized to play a role in C....
Show moreChlamydia trachomatis is an obligate, intracellular bacterium which is known to cause multiple human infections including nongonococcal urethritis (serovars D-K), lymphogranuloma venereum (serovars L1, L2, L3) and trachoma (serovars A-C). The infectious form of the bacterium, called the elementary body (EB), harbors a type III secreted effector known as Tarp (translocated actin recruiting phosphoprotein) which is a candidate virulence factor and is hypothesized to play a role in C. trachomatis' ability to invade and grow within epithelial cells in a human host. C. trachomatis L2 Tarp harbors five unique protein domains which include the Phosphorylation Domain, the Proline Rich Domain, the Actin Binding Domain, and two F-Actin Binding Domains. Tarp has been biochemically characterized in vitro, but it has yet to be characterized in vivo due to a lack of genetic tools in C. trachomatis. Through the recent generation of a chlamydial transformation system, we have created transformants which express epitope tagged wild type or mutant Tarp effectors. In this thesis, C. trachomatis transformants expressing Tarp lacking one of the five biochemically defined protein domains were used to examine both bacterial invasion and bacterial development within mammalian host cells. Our results demonstrate that those EBs which harbor mutant Tarp missing either its Phosphorylation Domain or its Actin Binding Domain were less capable of host cell invasion. However, these transformants, once internalized, were capable of normal development when compared to wild type C. trachomatis or C. trachomatis harboring an epitope tagged wild type Tarp effector. These results suggest that transformant expressed Tarp lacking the Phosphorylation Domain or Actin Binding Domain may be acting as a dominant-negative effector protein. Ultimately, these results support the hypothesis that Tarp is a virulence factor for Chlamydia trachomatis. Furthermore, this data indicates that through the manipulation of the Tarp effector, C. trachomatis pathogenesis may be attenuated.
Show less
Date Issued: 2016
Identifier: CFE0006159, ucf:51142
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006159

Title: Synthesis and Characterization of Core-Shell Zinc Silica Nanoparticles and Zinc Silica Nanogels for Agricultural Applications.
Creator: Berroth, Megan, Santra, Swadeshmukul, Moore, Sean, Jewett, Travis, University of Central Florida
Abstract / Description: Plant pathogens are a serious problem facing the agricultural industry today. Current methodologies use copper based biocides as the main form of defense. Unfortunately this can lead to damaging environmental effects and increased rates of antimicrobial resistance. In this study, antimicrobial activity of multiple alternative zinc-based nanoformulations were tested against three important plant pathogens: Xanthomonas alfalfae, Pseudomonas syringae, and Clavobacter michiganensis. Xanthomonas...
Show morePlant pathogens are a serious problem facing the agricultural industry today. Current methodologies use copper based biocides as the main form of defense. Unfortunately this can lead to damaging environmental effects and increased rates of antimicrobial resistance. In this study, antimicrobial activity of multiple alternative zinc-based nanoformulations were tested against three important plant pathogens: Xanthomonas alfalfae, Pseudomonas syringae, and Clavobacter michiganensis. Xanthomonas sub species cause Citrus canker, a devastating disease that affects millions of citrus trees worldwide while the latter two affect tomato crops. Materials synthesis was completed and the resulting nanoformulations were characterized by Atomic Absorption Spectroscopy, Scanning Electron Microscopy, High Resolution Transmission Electron Microscopy, and X-Ray Photoelectron Spectroscopy. The antimicrobial efficacy of the newly synthesized formulas and two commercially available products, Kocide 3000 (DuPont) and Nordox (Brandt), were determined by Minimum Inhibitory Concentration Assays followed by Bacterial Viability Assays. The subsequent data demonstrated a marketed difference in the way the antimicrobial agents acted upon the bacterial species. The core-shell zinc silica nanoparticles (C-SZnSiNP) proved to be ineffective, while the zinc silica nanogel (ZnSiNG) was as successful at killing the bacteria as the commercial products. This shows promise for a new alternative material with zinc at the forefront of the fight against plant pathogens.
Show less
Date Issued: 2015
Identifier: CFE0006209, ucf:51099
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006209

Title: Synthesis and Characterization of Antimicrobial Non-Color Forming Silica-Silver Nanocomposite.
Creator: Bazata, Joshua, Santra, Swadeshmukul, Moore, Sean, Jewett, Travis, University of Central Florida
Abstract / Description: Silver has been utilized for its antimicrobial properties for thousands of years in a variety of fields, extending the shelf life of food and water, rendering eating utensils sanitary, and more recently in biomedical applications such as silver based antiseptic creams. While effective as an antimicrobial agent at very low concentrations ((&)#181;g/mL), silver imparts a strong color to objects it is incorporated into, due to its high plasmonic efficiency. The goal of this study was to...
Show moreSilver has been utilized for its antimicrobial properties for thousands of years in a variety of fields, extending the shelf life of food and water, rendering eating utensils sanitary, and more recently in biomedical applications such as silver based antiseptic creams. While effective as an antimicrobial agent at very low concentrations ((&)#181;g/mL), silver imparts a strong color to objects it is incorporated into, due to its high plasmonic efficiency. The goal of this study was to determine if incorporating silver nanoparticles into a silica matrix could reduce or eliminate the plasmonic signal, while retaining the antimicrobial effects of the silver nanoparticles.Citrate capped silver nanoparticles (AgNP) were synthesized using a borohydride reduction method as outlined by Zheng et. al., and incorporated into silica nanoparticles using a method adapted from Fleger et. al. To test the antimicrobial efficacy of these synthesized silica coated silver nanoparticles (SiAgNP), minimum inhibitory concentration testing at three time points, 1, 4, and 8 hours, was carried out against E. coli and S. aureus using broth microdilution and Alamar Blue as an indicator of microbial growth. Efficacy was judged against uncoated AgNP and aqueous silver nitrate (AgNO3) solutions at equivalent Ag concentrations. Silica nanoparticles (SiNP) were utilized as a negative control. Further antimicrobial characterization using a bacterial viability assay revealed a time dependent killing trend in the SiAgNP, suggesting a controlled release of Ag+ from within the silica matrix. Efficacy of the SiAgNP was determined to fall between the most effective antimicrobial form of silver tested, AgNO3, and least effective, AgNP. However, the SiAgNP material exhibited no visible plasmon peak when UV-Visible spectrophotometric readings were taken, as well as remaining colorless when coated onto a ceramic substrate. Zeta potential revealed a high degree of colloidal stability of the SiAgNP. TEM imaging studies were carried out, verifying the presence of Ag within and on the silica nanoparticles, as well as the crystalline structure of the uncoated AgNP. It was determined that coating AgNP synthesized through borohydride reduction with silica through a St(&)#246;ber synthesis mechanism yields a material with enhanced antimicrobial effects compared to AgNP, but with no detectable plasmon signal, effectively producing a non-color forming silver based antimicrobial.
Show less
Date Issued: 2015
Identifier: CFE0006208, ucf:51097
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006208

Title: Host and Bacterial Determinants of Staphylococcus aureus Nasal Colonization in Humans.
Creator: Muthukrishnan, Gowrishankar, Cole, Alexander, Moore, Sean, Self, William, Parkinson, Christopher, University of Central Florida
Abstract / Description: Staphylococcus aureus (SA), an opportunistic pathogen colonizing the anterior nares in approximately 30% of the human population, causes severe hospital-associated and community-acquired infections. SA nasal carriage plays a critical role in the pathogenesis of staphylococcal infections and SA eradication from the nares has proven to be effective in reducing endogenous infections. To understand SA nasal colonization and its relation with consequent disease, assessment of nasal carriage...
Show moreStaphylococcus aureus (SA), an opportunistic pathogen colonizing the anterior nares in approximately 30% of the human population, causes severe hospital-associated and community-acquired infections. SA nasal carriage plays a critical role in the pathogenesis of staphylococcal infections and SA eradication from the nares has proven to be effective in reducing endogenous infections. To understand SA nasal colonization and its relation with consequent disease, assessment of nasal carriage dynamics among a diverse population and determining factors responsible for SA nasal carriage have become major imperatives.Here, we report on an extensive longitudinal monitoring of SA nasal carriage in 109 healthy individuals over a period of up to three years to assess nasal carriage dynamics. Phylogenetic analyses of SA housekeeping genes and hypervariable virulence genes revealed that not only were SA strains colonizing intermittent and persistent nasal carriers genetically similar, but no preferential colonization of specific SA strains in these carriers was observed over time. These results indicated that other non-SA factors could be involved in determining specific carriage states. Therefore, to elucidate host responses during SA nasal carriage, we performed human SA nasal recolonization in a subset of SA nasal carriers within our cohort. In these studies, SA colonization levels were determined, and nasal secretions were collected and analyzed for host immune factors responsible for SA nasal carriage. Interestingly, we observed that stimulation of host immune responses lead to clearance of SA while sustained SA colonization was observed in hosts that did not mount a response during carriage. Further, analysis of nasal secretions from hosts revealed that proinflammatory cytokines and chemokines were significantly induced during SA nasal clearance suggesting that innate immune effectors influence carriage.SA utilizes a repertoire of surface and secreted proteins to evade host immune response and successfully colonize the nose. Analysis of the most abundant immunoevasive proteins in the exoproteome of SA nasal carrier strains revealed that expression levels of Staphylococcal protein A (SPA) produced by SA nasal carrier strains in vitro corresponded to the level of persistence of SA in the human nose. To determine if SPA is involved in modulating the host's response to SA colonization, a subset of participants in our cohort was nasally recolonized with equal concentrations of both wild-type (WT) and spa-disrupted (?spa) autologous strains of SA. Interestingly, ?spa strains were eliminated from the nares significantly faster than WT when the host mounted an immune response, suggesting that the immunoevasive role of SPA is a determinant of carriage persistence. Collectively, this report augments our understanding of SA nasal carriage dynamics, in addition to identifying important host and microbial determinants that influence SA nasal colonization in humans. Better understanding of this phenomenon can lead to improved preventative strategies to thwart carriage-associated SA infections.
Show less
Date Issued: 2014
Identifier: CFE0005673, ucf:50173
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005673

Title: Screening of Quantum Dots for Toxicity on the Growth and Viability of Escherichia coli.
Creator: Tharkur, Jeremy, Santra, Swadeshmukul, Self, William, Moore, Sean, University of Central Florida
Abstract / Description: Heavy metal (HM) containing quantum dots (Qdots) are increasingly used in commercial products due to their unique electronic, optoelectronic, optical and magnetic properties. Once disposed to the landfill, environmental weathering is likely to compromise HM Qdot integrity, leading to release of heavy metal ions. To minimize any negative environmental impact of HM Qdots, there is an increasing demand for developing HM free or environmentally-friendly surface modified HM Qdot alternatives. In...
Show moreHeavy metal (HM) containing quantum dots (Qdots) are increasingly used in commercial products due to their unique electronic, optoelectronic, optical and magnetic properties. Once disposed to the landfill, environmental weathering is likely to compromise HM Qdot integrity, leading to release of heavy metal ions. To minimize any negative environmental impact of HM Qdots, there is an increasing demand for developing HM free or environmentally-friendly surface modified HM Qdot alternatives. In this study, synthesis of HM free ZnS:Mn/ZnS and surface modified HM CdS:Mn/ZnS Qdots (using N-acetylcysteine, NAC, and Dihydrolipoic acid, DHLA) and their potential toxicity assessment using E. coli as a model system is reported. NAC and DHLA are known antioxidants and therefore expected to reduce HM induced toxicity and improve colloidal stability of Qdots. All Qdots were synthesized at room temperature using a reverse micelle microemulsion method. Qdots were fully characterized using UV-visible absorption spectroscopy, fluorescence emission spectroscopy, zeta potential, Nuclear Magnetic Resonance spectroscopy (NMR) and High Resolution Transmission Electron Microscopy (HRTEM). Qdot environmental weathering was simulated by treating Qdots with concentrated acid (6N HCl). Qdot toxicity was evaluated on E. coli growth and viability using growth curves, turbidity and bactericidal assays (CFU). Results show that Zn based Qdots exhibit reduced toxicity on E.coli growth and viability when compared to Cd based Qdots. In addition, surface modification with NAC and DHLA minimized toxicity of Cd based Qdots. In summary, Zn based Qdots appear to be more environmental-friendly than Cd based Qdots.
Show less
Date Issued: 2013
Identifier: CFE0005426, ucf:50416
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005426

Title: Purification and Characterization of a Novel Selenocysteine Lyase from Enterococcus faecalis.
Creator: Nelson, Samantha, Self, William, Moore, Sean, Rohde, Kyle, University of Central Florida
Abstract / Description: A previous study identified Enterococcus faecalis as one of two bacteria known to have the selD gene and other selenium related genes without having the genes necessary to make selenocysteine or selenouridine. EF2570, a gene in the cluster, was later shown to be upregulated during biofilm formation and also responsible for a selenite- and molybdate-dependent increase in biofilm formation in vitro. The protein encoded was identified as a selenium dependent molybdenum hydroxylase (SDMH),...
Show moreA previous study identified Enterococcus faecalis as one of two bacteria known to have the selD gene and other selenium related genes without having the genes necessary to make selenocysteine or selenouridine. EF2570, a gene in the cluster, was later shown to be upregulated during biofilm formation and also responsible for a selenite- and molybdate-dependent increase in biofilm formation in vitro. The protein encoded was identified as a selenium dependent molybdenum hydroxylase (SDMH), enzymes that contain a labile selenium atom required for activity. While the process of inserting selenocysteine into a protein is well known, the process by which a SDMH acquires a labile selenium atom has not yet been described. To begin unraveling this pathway, the nifS-like EF2568 from the gene cluster will be characterized. Some NifS-like proteins have been shown to have selenocysteine lyase activity, providing a source of selenium for selenophosphate synthetase, the selD gene product. Study of EF2568 has shown that it specifically reacts with L-selenocysteine to form selenide and alanine with L-cysteine inhibiting the reaction. Guided by homology to the well-characterized human and E. coli NifS-like proteins, mutants of the active site and substrate discerning residues were also characterized for activity with L-selenocysteine and L-cysteine. While mutation of the residue at position 112 thought to be responsible for substrate specificity did not affect reactivity of the enzyme with L-cysteine, it did affect reactivity with L-selenocysteine. Studying the characteristics of this novel group II selenocysteine lyase will provide a foundation for studying the remaining pathway.
Show less
Date Issued: 2014
Identifier: CFE0005388, ucf:50455
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005388

Title: Mixed Valence Copper(Cu)/Silica Nanocomposite: Synthesis, Characterization and Systematic Antimicrobial Studies.
Creator: Young, Mikaeel, Santra, Swadeshmukul, Self, William, Moore, Sean, University of Central Florida
Abstract / Description: Copper (Cu) compounds are widely used as effective agricultural bactericides. Continuous use of these materials has led to Cu accumulation in soil over time. The United States Environmental Protection Agency (US EPA) is concerned about potential Cu contamination in the environment. Improving biocidal efficacy of Cu is an attractive alternative, allowing reduction of Cu amount per application. In this research, we focused on making water-soluble mixed-valence Copper/Silica composite nanogel ...
Show moreCopper (Cu) compounds are widely used as effective agricultural bactericides. Continuous use of these materials has led to Cu accumulation in soil over time. The United States Environmental Protection Agency (US EPA) is concerned about potential Cu contamination in the environment. Improving biocidal efficacy of Cu is an attractive alternative, allowing reduction of Cu amount per application. In this research, we focused on making water-soluble mixed-valence Copper/Silica composite nanogel (CuSiNG) material. The objective is to improve the efficacy of Cu by manipulating Cu valence states. It has been shown in the literature that Cu (0) and Cu (I) states are more potent that Cu (II) states in terms of their antimicrobial efficacy. It is hypothesized that mixed valence Cu will exhibit improved efficacy over Cu (II). A water-soluble mixed valence Cu/silica nanogel (MV-CuSiNG) composite has been synthesized and characterized. Structure, morphology, crystallinity and composition of the MV-CuSiNG material was characterized using High-Resolution Transmission Electron Microscopy (HRTEM), HRTEM Selected Area Electron Diffraction (SAED) and X-ray Photoelectron Spectroscopy (XPS). Amount of Cu loading in MV-CuSiNG composite material was estimated by Atomic Absorption Spectroscopy (AAS). To confirm presence of Cu (I) in the MV-CuSiNG material, Neocuproine (Nc, a Cu (I) specific chelator) assay was used. Antimicrobial efficacy of MV-CuSiNG and CuSiNG was evaluated against X.alfalfae, B.subtilis and E.coli using Kocide(&)#174; 3000 ((")Insoluble Cu (II)(") compound), Copper sulfate ((")Soluble Cu (II)(") compound) and Cuprous chloride (Copper (I) compound) as positive controls and silica (")seed(") particles (without Cu loading) as negative control. Antimicrobial studies included observing bacterial growth inhibition and determining the Minimum Inhibitory Concentration (MIC). Improved antimicrobial efficacy was observed in MV-CuSiNG when compared to CuSiNG and other controls. For the assessment of plant safety of MV-CuSiNG and CuSiNG materials, phytotoxicity studies were conducted using Vinca sp and Hamlin orange under environmental conditions. It was observed that MV-CuSiNG material was safe to plants at commercially used (standard) spray application rate.
Show less
Date Issued: 2013
Identifier: CFE0005282, ucf:50550
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005282

Title: Characterization of Novel Borrelia burgdorferi Transcripts Expressed during Tick and Mammalian Infection.
Creator: Adams, Philip, Jewett, Mollie, Rohde, Kyle, Moore, Sean, Fernandez-Valle, Cristina, University of Central Florida
Abstract / Description: The purpose of this dissertation is to characterize the transcriptome of Borrelia (Borreliella) burgdorferi to discover novel transcripts, important for pathogenesis. As a spirochete and the etiological agent of Lyme disease, the foremost vector-borne bacterial infection in the world, B. burgdorferi fulfills a distinctive niche among bacterial pathogens. Persisting in the disparate environments of a tick vector and mammalian reservoirs, it is absolutely dependent on its hosts for transmission...
Show moreThe purpose of this dissertation is to characterize the transcriptome of Borrelia (Borreliella) burgdorferi to discover novel transcripts, important for pathogenesis. As a spirochete and the etiological agent of Lyme disease, the foremost vector-borne bacterial infection in the world, B. burgdorferi fulfills a distinctive niche among bacterial pathogens. Persisting in the disparate environments of a tick vector and mammalian reservoirs, it is absolutely dependent on its hosts for transmission and nutrient acquisition. B. burgdorferi harbors a complex fragmented genome which is largely linear, unlike that of most prokaryotes, lacks an array of classically described metabolic genes, and contains an unusually large percentage of unique genomic sequences specific to Borrelia (Borreliella) species. To date, few regulatory mechanisms have been identified which contribute to the ability of the spirochete to sense and respond to its environment. Efforts to use global transcript analysis to elucidate the molecular mechanisms of B. burgdorferi host adaptation have proven challenging due to the low numbers of the pathogen present during infection. Previously, our laboratory successfully developed an in vivo expression technology based approach for B. burgdorferi (BbIVET) to identify spirochete promoter sequences that are active during a murine infection. This screen identified 233 unique putative promoters which mapped to locations across the entire genome. These putative infection-active B. burgdorferi promoters were not only located at the 5' end of annotated open reading frames (ORFs), but also mapped to unannotated locations antisense, intergenic, and intragenic to ORFs. Given the limited characterization of the B. burgdorferi transcriptome, this dissertation applies an RNA sequencing approach (5'RNA-seq) to globally annotate the transcriptional start sites (TSSs) and 5' processed ends of the spirochete's RNA during in vitro cultivation. This resulted in the discovery of numerous novel internal, intergenic, and antisense transcripts. Synergistic analysis combining Northern blotting techniques, alignments of these transcripts to BbIVET proposed promoters, and interrogation of promoter activity via in vivo live imaging of mice, confirmed the expression of a variety of RNAs during laboratory culture and mammalian infection. Further, as a means to improve quantitation of the expression of these transcripts, a new methodology was developed and applied to measure B. burgdorferi promoter activity during tick-pathogen interactions, in a strand specific manner. Finally, because the Lyme disease spirochete harbors many unclassified and unique genomic sequences, the mammalian infection-expressed gene bb0562, identified through BbIVET and 5'RNA-seq, was selected for targeted deletion and evaluation throughout B. burgdorferi's infectious cycle. This demonstrated that gene bb0562 encodes a membrane associated protein, whose presence is critical for establishing murine infection through the bite of an infected tick. In sum, this work contributes significant insight into the transcriptome of B. burgdorferi, provides an innovative approach for the analysis of RNA transcripts at the tick-pathogen interface, and identifies a novel gene critical for Lyme disease pathogenesis.
Show less
Date Issued: 2017
Identifier: CFE0006707, ucf:51915
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006707

Title: An Assessment of Biosorption Activated Media for the Removal of Pollutants in Up-Flow Stormwater Treatment Systems.
Creator: Hood, Andrew, Randall, Andrew, Wanielista, Martin, Chopra, Manoj, O'Reilly, Andrew, Moore, Sean, University of Central Florida
Abstract / Description: Nitrogen and phosphorus are often the limiting nutrients for marine and freshwater systems respectively. Additionally, stormwater often contains elevated levels of pathogens which can pollute the receiving water body and impact reuse applications [1-4]. The reduction of limiting nutrients and pathogens is a common primary target for stormwater best management practices (BMPs) [5]. Traditional BMPs, such as retention/detention treatment ponds require large footprints and may not be practical...
Show moreNitrogen and phosphorus are often the limiting nutrients for marine and freshwater systems respectively. Additionally, stormwater often contains elevated levels of pathogens which can pollute the receiving water body and impact reuse applications [1-4]. The reduction of limiting nutrients and pathogens is a common primary target for stormwater best management practices (BMPs) [5]. Traditional BMPs, such as retention/detention treatment ponds require large footprints and may not be practical in ultra-urban environments where above ground space is limited. Upflow filters utilizing biosorption activated media (BAM) that can be placed underground offer a small footprint alternative. Additionally, BAM upflow filters can be installed at the discharge point of traditional stormwater ponds to provide further treatment. This research simulated stormwater that had already been treated for solids removal; thus, most of the nutrients and solids in the influent were assumed to be as non-settable suspended solids or dissolved solids. Three different BAM mixtures in an upflow filter configuration were compared for the parameters of nitrogen, phosphorus, total coliform, E. coli, and heterotrophic plate count (HPC). Additionally, genetic testing was conducted using Polymerase Chain Reaction (PCR), in conjunction with a nitrogen mass balance, to determine if Anammox was a significant player in the nitrogen removal. The columns were run at both 22-minute and 220-minute Empty Bed Contact Times (EBCTs). All the BAM mixtures analyzed were shown to be capable at the removal of nitrogen, phosphorus, and total coliform during both the 22-minute and 220-minute EBCTs, with BAM #1 having the highest removal performance for all three parameters during both EBCTs. All BAM mixtures experienced an increase in HPC. Additionally, PCR analysis confirmed the presence of Anammox in the biofilm and via mass balance it was determined that the biological nitrogen removal was due to Anammox and endogenous denitrification with Anammox being a significant mechanism.
Show less
Date Issued: 2019
Identifier: CFE0007817, ucf:52875
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007817

Title: Investigating Novel Water Treatment Methods and Monitoring Techniques for Sulfide-Laden Groundwater Supplies.
Creator: Yoakum, Benjamin, Duranceau, Steven, Lee, Woo Hyoung, Sadmani, A H M Anwar, Moore, Sean, University of Central Florida
Abstract / Description: This dissertation reports on research related to novel water treatment and monitoring techniques for sulfide-laden groundwater supplies. The dissertation is divided into several chapters with four core chapters focused on investigations studying a novel water treatment method or monitoring technique. The first investigation assessed the efficacy of multi-pass spray aeration treatment to remove trihalomethanes (THMs) and to reduce the total THM formation potential (TTHMFP) of an aerated water...
Show moreThis dissertation reports on research related to novel water treatment and monitoring techniques for sulfide-laden groundwater supplies. The dissertation is divided into several chapters with four core chapters focused on investigations studying a novel water treatment method or monitoring technique. The first investigation assessed the efficacy of multi-pass spray aeration treatment to remove trihalomethanes (THMs) and to reduce the total THM formation potential (TTHMFP) of an aerated water column post-aeration. A recirculating spray aeration pilot unit was constructed to make this assessment. To assess the effect of multi-pass spray aeration on the TTHMFP, water was recirculated through a fabricated spray nozzle for various lengths of time. Results showed that multi-pass spray aeration can remove chloroform, dichlorobromomethane, dibromochloromethane and bromoform to below detection levels ((<) 0.7 ppb) for the waters investigated. Additionally, spray aeration reduced the TTHMFP of chlorinated water. Results suggest multi-pass spray aeration may be a viable treatment option for some bromide container waters. Results also indicate that multi-pass spray aeration removes bromide from the bulk water in the form of organically bound volatile compounds.The second investigation assessed the efficacy of using pre-existing tray aeration infrastructure to comply with disinfection by-product (DBP) regulations. To assess the efficacy of tray aerators to reduce the concentration TTHMs a pilot tray aerator was constructed. Results showed that after five tray passes (each pass consisting of water being passed over five trays) the concentration of TTHMs was below the detection limit ((<) 0.7 ppb) for the water investigated. To assess the efficacy of tray aeration at full-scale, a water treatment plant and the distribution system it serves were monitored for eight months. Results showed an approximate 40 ppb reduction in the TTHM concentration at two on-site monitoring locations and the one off-site monitoring location (initial concentrations being approximately 54 ppb, 60 ppb and 73 ppb, respectively). Results suggest that the utility managing the full-scale system could comply with DBP regulations by using the pre-existing tray aeration infrastructure to reduce formed THMs on-site where regulated haloacetic acids are not predominant.The third investigation assessed the efficacy of using biological activated carbon (BAC) to remove disinfection by-product precursor matter to comply with DBP regulations. To research this method, a pilot scale BAC filter was operated for three independent test runs. In addition, two full-scale WTPs using BAC were monitored over time. Results showed an approximate 40 percent removal of dissolved organic carbon (DOC) during the three pilot runs and an approximate 55 percent removal of DOC during full-scale monitoring. Results showed that the reduction in DOC reduced the TTHMFP of BAC treated water. Results suggest that BAC treatment could be a viable treatment option to comply with DBP regulations in the sulfide-laden water studied.The fourth investigation assessed the suitability of oxidation reduction potential (ORP) to monitor the effectiveness of an oxidizing media filter used to remove sulfur from a sulfide-laden groundwater. Results showed that ORP was more useful as a measurement technique as compared to free chlorine residual when assessing filter bed health and regeneration effectiveness. It was determined that when the ORP measurement taken from within the oxidative media layer was below 500 mV, the filter bed was not providing treatment, and manganese could be released. Results showed a significant increase in turbidity ((>) 2 NTU) and total manganese ((>) 0.05 mg/L) occurred when the ORP within the filter bed dropped below 400 mV. More frequent cycling of the filters was found to be an effective treatment option to maintain ORP values above an identified 400 mV operational threshold.
Show less
Date Issued: 2017
Identifier: CFE0007141, ucf:52317
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007141

Title: Multi-target high-throughput screening assays for antimicrobial drug discovery.
Creator: Grube, Christopher, Roy, Herve, Chakrabarti, Debopam, Moore, Sean, Koculi, Eda, University of Central Florida
Abstract / Description: The rise of antibiotic resistant microbes (bacteria, fungi, and parasites), combined with the current void of new drugs entering the clinical setting, has created an urgent need for the discovery of new antimicrobials. High-throughput screening (HTS) assays represent a fast and cost-efficient method for identifying new therapeutic compounds and have been the longstanding gold standard for drug discovery. The focus of this dissertation is on the development and implementation of novel...
Show moreThe rise of antibiotic resistant microbes (bacteria, fungi, and parasites), combined with the current void of new drugs entering the clinical setting, has created an urgent need for the discovery of new antimicrobials. High-throughput screening (HTS) assays represent a fast and cost-efficient method for identifying new therapeutic compounds and have been the longstanding gold standard for drug discovery. The focus of this dissertation is on the development and implementation of novel methodologies to increase the throughput of target-based HTS by designing assays that allow multiple drug targets to be probed simultaneously. During my graduate studies, I developed three distinct HTS assays. In each of these assays, drug targets were incorporated into synthetic pathways obeying various reaction topologies (e.g., cyclical, parallel, or linear). Each of these reaction topologies conferred specific advantages and limitations to the individual assays. The first assay reconstitutes the bacterial tRNA-dependent pathway for lipid aminoacylation. This two-step pathway combines a tRNA aminoacylation step catalyzed by an aminoacyl-tRNA synthetase (aaRS), and a transferase step, which transfers the amino acid born by the tRNA onto membrane lipids. aaRSs are essential enzymes in all domains of life and represent longstanding drug targets in pathogenic species. The transferase reaction in the pathway is also an appealing drug target since it impacts the cellular permeability of antibiotics. Inhibitors of this reaction could dramatically increase the efficacy of existing therapeutics. The second assay I developed also targets aaRSs, but utilizes a parallel topology that permits the probing of the synthetic and editing activities of up to four aaRSs simultaneously. The third assay utilizes a linear topology that reconstitutes the entire purine salvage pathway from Plasmodium falciparum. Because parasites are unable to synthesize purines de novo, this pathway represents an appealing target for novel antimalarials. Pilot screens using this assay revealed inhibitors for multiple enzymes in the pathway, validating the design of the system. This body of work aims to shift the current paradigm of single-target systems that have historically dominated the HTS field, toward multi-target designs that can be used to more efficiently screen compound libraries against essential pathways in pathogenic microbes.
Show less
Date Issued: 2019
Identifier: CFE0007642, ucf:52469
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007642

Title: Machine Learning Methods for Multiparameter Flow Cytometry Analysis and Visualization.
Creator: Sassano, Emily, Jha, Sumit Kumar, Pattanaik, Sumanta, Hughes, Charles, Moore, Sean, University of Central Florida
Abstract / Description: Flow cytometry is a popular analytical cell-biology instrument that uses specific wavelengths of light to profile heterogeneous populations of cells at the individual level. Current cytometers have the capability of analyzing up to 20 parameters on over a million cells, but despite the complexity of these datasets, a typical workflow relies on subjective labor-intensive manual sequential analysis. The research presented in this dissertation provides two machine learning methods to increase...
Show moreFlow cytometry is a popular analytical cell-biology instrument that uses specific wavelengths of light to profile heterogeneous populations of cells at the individual level. Current cytometers have the capability of analyzing up to 20 parameters on over a million cells, but despite the complexity of these datasets, a typical workflow relies on subjective labor-intensive manual sequential analysis. The research presented in this dissertation provides two machine learning methods to increase the objectivity, efficiency, and discovery in flow cytometry data analysis. The first, a supervised learning method, utilizes previously analyzed data to evaluate new flow cytometry files containing similar parameters. The probability distribution of each dimension in a file is matched to each related dimension of a reference file through color indexing and histogram intersection methods. Once a similar reference file is selected the cell populations previously classified are used to create a tailored support vector machine capable of classifying cell populations as an expert would. This method has produced results highly correlated with manual sequential analysis, providing an efficient alternative for analyzing a large number of samples. The second, a novel unsupervised method, is used to explore and visualize single-cell data in an objective manner. To accomplish this, a hypergraph sampling method was created to preserve rare events within the flow data before divisively clustering the sampled data using singular value decomposition. The unsampled data is added to the discovered set of clusters using a support vector machine classifier, and the final analysis is displayed as a minimum spanning tree. This tree is capable of distinguishing rare subsets of cells comprising of less than 1% of the original data.
Show less
Date Issued: 2018
Identifier: CFE0007243, ucf:52241
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007243

Infectious disease (3)	+ -
Antimicrobial (2)	+ -
E. coli (2)	+ -
Enterococcus faecalis (2)	+ -
Nasal Colonization (2)	+ -

Phylogenetics (2)	+ -
Silica (2)	+ -
cysteine desulfurase (2)	+ -
ribosome (2)	+ -
selenocysteine lyase (2)	+ -
translation (2)	+ -
16S rRNA gene sequencing (1)	+ -
5' end transcriptome (1)	+ -
ABC (1)	+ -
ABCE1 (1)	+ -
ATPase (1)	+ -
Aminoacyl-tRNA synthetase (1)	+ -
Ammonification (1)	+ -
Anammox (1)	+ -
BAM (1)	+ -
BMP (1)	+ -
Best Management Practice (1)	+ -
Biocide (1)	+ -
Biological Filtration (1)	+ -
Biological Treatment (1)	+ -
Biosorption (1)	+ -
Biosorption Activated Media (1)	+ -
Bold & Gold (1)	+ -
Bold and Gold (1)	+ -
Borrelia (Borreliella) burgdorferi (1)	+ -

ETD Collection (23)	+ -
Honors Collection (3)	+ -

University of Central Florida (26)	+ -
Moore, Sean (25)	+ -
Self, William (7)	+ -
Roy, Herve (6)	+ -
Cole, Alexander (5)	+ -

Santra, Swadeshmukul (4)	+ -
Jewett, Travis (3)	+ -
Teter, Kenneth (3)	+ -
Harper, James (2)	+ -
Koculi, Eda (2)	+ -
Parkinson, Christopher (2)	+ -
Rohde, Kyle (2)	+ -
Adams, Philip (1)	+ -
Almagro-Moreno, Salvador (1)	+ -
Bazata, Joshua (1)	+ -
Berroth, Megan (1)	+ -
Boote, David (1)	+ -
Butler, Malcolm (1)	+ -
Campiglia, Andres (1)	+ -
Chai, Xinqing (1)	+ -
Chakrabarti, Debopam (1)	+ -
Chini, Jacquelyn (1)	+ -
Chopra, Manoj (1)	+ -
Chumbimuni Torres, Karin (1)	+ -
Costello, Michael S (1)	+ -
Dawkins, Molly C (1)	+ -
Duranceau, Steven (1)	+ -
Ebert, Steven (1)	+ -
Elsheimer, Seth (1)	+ -
Fedorka, Kenneth (1)	+ -

Search In

Pages

Pages

Sort Results By:

Narrow results by:

Topical Subject

Collection

Type of Resource

Language

Creator

Format

Owner