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(1 - 7 of 7)
- Title
- MAGNETIC NANOSENSORS FOR MULTIPLEXED BACTERIAL PATHOGENESIS IDENTIFICATION.
- Creator
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Kaittanis, Charalambos, Perez, J. Manuel, University of Central Florida
- Abstract / Description
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Developing diagnostic modalities that utilize nanomaterials and miniaturized detectors can have an impact in point-of-care diagnostics. Diagnostic systems that (i) are sensitive, robust, and portable, (ii) allow detection in clinical samples, (iii) require minimal sample preparation yielding results quickly, and (iv) can simultaneously quantify multiple targets, would have a great potential in biomedical research and public healthcare. Bacterial infections still cause pathogenesis throughout...
Show moreDeveloping diagnostic modalities that utilize nanomaterials and miniaturized detectors can have an impact in point-of-care diagnostics. Diagnostic systems that (i) are sensitive, robust, and portable, (ii) allow detection in clinical samples, (iii) require minimal sample preparation yielding results quickly, and (iv) can simultaneously quantify multiple targets, would have a great potential in biomedical research and public healthcare. Bacterial infections still cause pathogenesis throughout the world (Chapter I). The emergence of multi-drug resistant strains, the potential appearance of bacterial pandemics, the increased occurrence of bacterial nosocomial infections, the wide-scale food poisoning incidents and the use of bacteria in biowarfare highlight the need for designing novel bacterial-sensing modalities. Among the most prominent disease-causing bacteria are strains of Escherichia coli, like the E. coli O157:H7 that produces the Shiga-like toxin (Stx). Apart from diarrheagenic E. coli strains, others cause disease varying from hemolytic uremic syndrome and urinary tract infections to septicemia and meningitis. Therefore, the detection of E. coli needs to be performed fast and reliably in diverse environmental and clinical samples. Similarly, Mycobacterium avium spp. paratuberculosis (MAP), a fastidious microorganism that causes JohneÂÂ's disease in cattle and has been implicated in CrohnÂÂ's disease (CD) etiology, is found in products from infected animals and clinical samples from CD patients, making MAP an excellent proof-of-principle model. Recently, magnetic relaxation nanosensors (MRnS) provided the first applications of improved diagnostics with high sensitivity and specificity. Nucleic acids, proteins, viruses and enzymatic activity were probed, yet neither large targets (for instance bacterial and mammalian cells) nor multiple bacterial disease parameters have been simultaneously monitored, in order to provide thorough information for clinical decision making. Therefore, the goal of this study was to utilize MRnS for the sensitive identification of multiple targets associated with bacterial pathogenesis, while monitoring virulence factors at the microorganism, nucleic acid and virulence factor levels, to facilitate improved diagnosis and optimal treatment regimes. To demonstrate the versatility of MRnS, we used MAP as our model system, as well as several other pathogens and eukaryotic cell lines. In initial studies, we developed MRnS suitable for biomedical applications (Chapter II). The resulting MRnS were composed of an iron oxide core, which was caged within a biodegradable polymeric coating that could be further functionalized for the attachment of molecular probes. We demonstrated that depending on the polymer used the physical and chemical properties of the MRnS can be tailored. Furthermore, we investigated the role of polymer in the enzyme-mimicking activity of MRnS, which may lead to the development of optimized colorimetric in vitro diagnostic systems such as immunoassays and small-molecule-based screening platforms. Additionally, via facile conjugation chemistries, we prepared bacterium-specific MRnS for the detection of nucleic acid signatures (Chapter III). Considering that MAP DNA can be detected in clinical samples and isolates from CD patients via laborious isolation and amplification procedures requiring several days, MRnS detected MAPÂÂ's IS900 nucleic acid marker up to a single MAP genome copy detection within 30 minutes. Furthermore, these MRnS achieved equally fast IS900 detection even in crude DNA extracts, outperforming the gold standard diagnostic method of nested Polymerase Chain Reaction (nPCR). Likewise, the MRnS detected IS900 with unprecedented sensitivity and specificity in clinical isolates obtained from blood and biopsies of CD patients, indicating the clinical utility of these nanosensors. Subsequently, we designed MRnS for the detection of MAP via surface-marker recognition in complex matrices (Chapter III). Milk and blood samples containing various concentrations of MAP were screened and quantified without any processing via MRnS, obtaining dynamic concentration-dependent curves within an hour. The MAP MRnS were able not only to identify their target in the presence of interferences from other Gram positive and Gram negative bacteria, but could differentiate MAP among other mycobacteria including Mycobacterium tuberculosis. In addition, detection of MAP was performed in clinical isolates from CD patients and homogenized tissues from JohneÂÂ's disease cattle, demonstrating for the first time the rapid identification of bacteria in produce, as well as clinical and environmental samples. However, comparing the unique MAP quantification patterns with literature-available trends of other targets, we were prompted to elucidate the underlying mechanism of this novel behavior (Chapter IV). We hypothesized that the nanoparticle valency – the amount of probe on the surface of the MRnS – may have modulated the changes in the relaxation times (ΔΤ2) upon MRnS – target association. To address this, we prepared MAP MRnS with high and low anti-MAP antibody levels using the same nanoparticle formulation. Results corroborated our hypothesis, but to further bolster it we investigated if this behavior is target-size-independent. Hence utilizing small-molecule- and antibody-carrying MRnS, we detected cancer cells in blood, observing similar detection patterns that resembled those of the bacterial studies. Notably, a single cancer cell was identified via high-valency small-molecule MRnS, having grave importance in cancer diagnostics because a single cancer cell progenitor in circulation can effectively initiate the metastatic process. Apart from cells, we also detected the Cholera Toxin B subunit with valencly-engineered MRnS, observing similar to the cellular targetsÂÂ' diagnostic profiling behavior. Finally, as bacterial drug resistance is of grave healthcare importance, we utilized MRnS for the assessment of bacterial metabolism and drug susceptibility (Chapter V). Contrary to spectophotometric and visual nanosensors, their magnetic counterparts were able to quickly assess bacterial carbohydrate uptake and sensitivity to antibiotics even in blood. Two MRnS-based assay formats were devised relying on either the Concanavalin A (Con A)-induced clustering of polysaccharide-coated nanoparticles or the association between free carbohydrates and Con A-carrying MRnS. Overall, taking together these results, as well as those on pathogen detection and the recent instrumentation advancements, the use of MRnS in the clinic, the lab and the field should be anticipated.
Show less - Date Issued
- 2010
- Identifier
- CFE0002982, ucf:47954
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0002982
- Title
- A Multisystem Approach for the Characterization of Bacteria for Sustainable Agriculture.
- Creator
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Lee, Briana, Tetard, Laurene, Kang, Hyeran, Mason, Chase, University of Central Florida
- Abstract / Description
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The chemical, physical, and biological properties of bacteria developing resistance have been explored in animal based bacteria while plant bacteria have been largely neglected. Thus, the ability to probe changes in stiffness, adhesion, binding interactions and molecular traits of bacteria causing plant diseases is of great interest to develop a new generation of more potent, yet sustainable, pesticides. Our study aims to investigate the physical and chemical properties of bacterial systems,...
Show moreThe chemical, physical, and biological properties of bacteria developing resistance have been explored in animal based bacteria while plant bacteria have been largely neglected. Thus, the ability to probe changes in stiffness, adhesion, binding interactions and molecular traits of bacteria causing plant diseases is of great interest to develop a new generation of more potent, yet sustainable, pesticides. Our study aims to investigate the physical and chemical properties of bacterial systems, in particular their cell walls. Building upon this fundamental understanding of the cells, we also investigate the physicochemical responses associated to multivalent nanoparticle-based bactericide treatments on bacterial systems identified as pathogens in plant diseases. Here our efforts focus on developing a protocol for the fundamental understanding of Xanthomonas perforans, a strain known for causing bacterial spot in tomatoes and causing close to 50% losses in production. To support the design and accelerate the development of pesticides and treatments against this disease, we evaluate the changes bacteria undergo in the presence of the treatment. Using a silica nanoparticle-based treatment designed with a shell containing multivalent copper and quaternary ammonium, we compare bacteria pre- and post-treatment with infrared spectroscopy, atomic force microscopy (AFM)-based techniques, and TIRF microscopy. Statistical data analysis enables the identification of attributes that can potentially serve as markers to track the bacterial responses to the treatment in the future. Finally, we will discuss the exciting implications of this work, such as potential clues for the development of more potent treatments for resistant bacteria.
Show less - Date Issued
- 2018
- Identifier
- CFE0007038, ucf:52005
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0007038
- Title
- AEROMONAS HYDROPHILA IN AMPHIBIANS: HARMLESS BYSTANDER OR OPPORTUNISTIC PATHOGEN.
- Creator
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Rivas, Zachary P, Savage, Anna Evangeline, University of Central Florida
- Abstract / Description
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For several decades amphibian populations have been declining. Historically, the bacterium A. hydrophila (Ah) was hypothesized to be the causal factor in amphibian disease and population declines. However, with the discovery of a chytrid fungus, Batrachochytrium dendrobatidis (Bd) in 1998, which was identified on the skin of amphibians during documented mortality events, Ah research became of minor interest as focus shifted to Bd. Recent studies into the immunocompromising abilities of Bd,...
Show moreFor several decades amphibian populations have been declining. Historically, the bacterium A. hydrophila (Ah) was hypothesized to be the causal factor in amphibian disease and population declines. However, with the discovery of a chytrid fungus, Batrachochytrium dendrobatidis (Bd) in 1998, which was identified on the skin of amphibians during documented mortality events, Ah research became of minor interest as focus shifted to Bd. Recent studies into the immunocompromising abilities of Bd, however, have opened new questions about its relationship with Ah and their combined effects on a host. In this study, I explore the relationship between infection with these two pathogens, Bd and Ah, in two amphibian species from distinct regions of the United States. I developed a novel qPCR assay to measure the microbial load of Ah on the skin of two anuran species, Lithobates yavapaiensis (N=232) and Pseudacris ornata (N=169), which have confirmed Bd infections. I use a logistic regression model to identify whether significant relationships exist between these two pathogens, disease, and death. I find that even amongst the most severely infected frogs, Ah is not detectable on the skin and only appears post-mortem. I therefore conclude that Ah is an opportunistic bacterial pathogen, scavenging on anurans only after mortality events. This research is the first known study to quantitatively assess Ah in amphibians in conjunction with Bd. While there is no causal relationship between these pathogens, future work will examine potential Ah infections in other organs to more fully understand the relationship between Bd and Ah.
Show less - Date Issued
- 2016
- Identifier
- CFH2000013, ucf:45589
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFH2000013
- Title
- Allelic characterization and novel functions of the outer membrane porin U in Vibrio cholerae.
- Creator
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Sakib, Sk Nazmus, Almagro-Moreno, Salvador, Moore, Sean, Roy, Herve, University of Central Florida
- Abstract / Description
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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
- THE GLYCINE AND PROLINE REDUCTASE SYSTEMS: AN EVOLUTIONARY PERSPECTIVE AND PRESCENCE IN ENTEROBACTERIACEAE.
- Creator
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Witt, Joshua, Self, William, University of Central Florida
- Abstract / Description
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The Glycine and Proline Reduction systems are two of the best characterized selenoenzymes in bacteria and have been found to occur in a wide variety of clostridia . These enzymes are utilized to reduce glycine or D-proline to obtain energy via substrate level phosporylation or membrane gradients, respectively [6, 7]. This includes the pathogens C. difficile and C. botulinum [5, 8]. Strains of C. difficile are activate toxigenic pathways whenever either of these pathways is active within the...
Show moreThe Glycine and Proline Reduction systems are two of the best characterized selenoenzymes in bacteria and have been found to occur in a wide variety of clostridia . These enzymes are utilized to reduce glycine or D-proline to obtain energy via substrate level phosporylation or membrane gradients, respectively [6, 7]. This includes the pathogens C. difficile and C. botulinum [5, 8]. Strains of C. difficile are activate toxigenic pathways whenever either of these pathways is active within the cell [5, 8]. Though evolutionary studies have been conducted on ammonia producing bacteria none has been done to directly characterize these two system by themselves. This includes an understanding of whether or not this system is transferred between organisms, as many of the clostridia that are to be studied are known to have an "open genome." [8, 10] With this information we were able to generate a phylogenic model of the proline and glycine reduction systems. Through this analysis, we were able to account for many clostridial organisms that contain the system, but also many other organisms as well. These included enterobacteriaceae including a strain of the model organism, Escherichia coli. It was further concluded that Glycine Reductase was a much less centralized system and included a wide range of taxa while Proline Reductase was much more centralized to being within the phyla of firmicutes. It was also concluded that the strain of E. coli has a fully functional operon for Glycine Reductase.
Show less - Date Issued
- 2013
- Identifier
- CFH0004506, ucf:45149
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFH0004506
- Title
- Modeling Disease Impact of Vibrio-Phage Interactions.
- Creator
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Botelho, Christopher, Shuai, Zhisheng, Nevai, A, Zhang, Teng, Teter, Kenneth, University of Central Florida
- Abstract / Description
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Since the work of John Snow, scientists and medical professionals have understood that individuals develop cholera by means of consuming contaminated water. Despite the knowledge(&)nbsp;of cholera's route of infection, many countries have experienced and still experience endemic cholera. Cholera is caused by the Vibrio cholerae (V. cholerae) bacterium and presents with acute diarrhea and vomiting. If untreated, infected individuals may die due to dehydration. Cholera is a disease that most...
Show moreSince the work of John Snow, scientists and medical professionals have understood that individuals develop cholera by means of consuming contaminated water. Despite the knowledge(&)nbsp;of cholera's route of infection, many countries have experienced and still experience endemic cholera. Cholera is caused by the Vibrio cholerae (V. cholerae) bacterium and presents with acute diarrhea and vomiting. If untreated, infected individuals may die due to dehydration. Cholera is a disease that most commonly affects countries with poor infrastructure and water sanitation. Despite efforts to control cholera in such countries, the disease persists. One such example is Haiti which has been experiencing a cholera outbreak since 2010. While there has been much research in the field of microbiology to understand V. cholerae, there has been comparably less research in the field of mathematical biology to understand the dynamics of V. cholerae in the environment. A mathematical model of V. cholerae incorporating a phage population is coupled with a SIRS disease model to examine the impact of vibrio and phage interaction. It is shown that there might exist two endemic equilibria, besides the disease free equilibrium: one in which phage persist in the environment and one in which the phage fail to persist. Existence and stability of these equilibria are established. Disease control strategies based on vibrio and phage interactions are discussed.
Show less - Date Issued
- 2019
- Identifier
- CFE0007604, ucf:52544
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0007604
- Title
- An Assessment of Biosorption Activated Media for the Removal of Pollutants in Up-Flow Stormwater Treatment Systems.
- Creator
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Hood, Andrew, Randall, Andrew, Wanielista, Martin, Chopra, Manoj, O'Reilly, Andrew, Moore, Sean, University of Central Florida
- Abstract / Description
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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
