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Clam (Corbicula Fluminea) As A Potential Sentinel Of Human Norovirus Contamination In Freshwater, Xunyan Ye May 2012

Clam (Corbicula Fluminea) As A Potential Sentinel Of Human Norovirus Contamination In Freshwater, Xunyan Ye

Dissertations

The purpose of this study was to evaluate and validate the use of the clam Corbicula fluminea as a sentinel of human noroviruses (HuNoV) contamination in freshwater. The first specific aim was to develop a new method to extract HuNoV RNA from contaminated bivalves (e.g. oysters, clams) that would be much faster than existing methods. The procedure developed includes an initial total RNA extraction using TRI Reagent, followed by HuNoV RNA concentration and purification using biotinylated probe-capture technology. HuNoV RNA is finally detected by real-time RT-PCR. Using bivalve homogenates spiked with HuNoV, 100 PCR detection units of the virus ...


Genetic Variation In Potentially Virulent Vibrio Parahaemolyticus From The Northern Gulf Of Mexico, Nicholas Felix Noriea Iii May 2012

Genetic Variation In Potentially Virulent Vibrio Parahaemolyticus From The Northern Gulf Of Mexico, Nicholas Felix Noriea Iii

Dissertations

Vibrio parahaemolyticus (Vp) is a gram-negative bacterium found naturally in marine and estuarine environments. Vp is found in oysters including those which are later consumed by the public. Sub-populations of potentially virulent Vp contain specific virulence factors and are relevant human pathogens capable of causing gastroenteritis, wound infection, and death. The tdh and trh genes, both encoding hemolysins, have been correlated with the majority of clinical Vp isolates but have not been shown to be the definitive virulence factors.

A total of 146 Vp isolates from the northern Gulf of Mexico were collected and probed for ...


Effects Of Hypoxia And 4-Tert-Octylphenol On Gene Expression Profiles Of The Sheepshead Minnow (Cyprinodon Variegatus), Arthur Alan Karels May 2012

Effects Of Hypoxia And 4-Tert-Octylphenol On Gene Expression Profiles Of The Sheepshead Minnow (Cyprinodon Variegatus), Arthur Alan Karels

Dissertations

Hypoxia occurs in estuaries of northern Gulf of Mexico and world-wide, with increasing frequency/severity via eutrophication and anthropogenic influences. Hypoxia inducible factors (HIFs) form transcriptional complex and bind DNA at hypoxia responsive elements (HREs) in promoter regions of genes needed for systemic and cellular adaptation of fish to low dissolved oxygen (hypoxia, DO <2.0 mg/ml). Hypoxia-induced activation of HIF-αs can lead to a cascade of downstream activation, such as erythropoietin (EPO). Return to normal DO levels (normoxia), prolyl hydroxylases (PHDs) are activated to degrade HIF-αs back to baseline. Fish are affected by environmental estrogen mimics, like 4-tert-octylphenol (4tOP), binding estrogen receptor alpha (ERα) at estrogen responsive elements (EREs) and activating genes vitellogenin (VTG). Previous research showed overlap or crosstalk between these two mechanistic pathways. Hypoxia triggers unknown factors regulating ERE-mediated ERα signaling pathway, and stressor combinations could increase/decrease hypoxic or endocrine pathway. Research examined molecular/physiological effects of hypoxia (acute and chronic, moderate and severe) and 4tOP (~60μg/L)on adult male and/or female sheepshead minnow (Cyprinodon variegatus). Three genes identified, cloned, and sequenced (HIF-1α, HIF-2α, and PHD3), plus previously identified genes EPO and VTG, were examined in liver/testes exposed to hypoxia and/or 4tOP for cellular/physiological changes. Endpoints examined included mRNA expression from real-time PCR of HIF-1α, HIF-2α, PHD3, EPO, and VTG using cDNA from total RNA extracts, and microarray analyses of genes expressed during the transition from hypoxia back to normoxia. Phylogenetic analyses confirmed isolation of two HIF-α isofoms (HIF-1α and HIF-2α) and the PHD3 isoform. Significant up-regulation of PHD3 occurred within 10 hrs of chronic hypoxia, and persisted when severe (1.5 mg/L) and declined when moderate (~2.5mg/L). Significant up-regulation of HIF-1α and EPO occurred within 30 minutes to 2 hours of onset of acute severe and very severe (~1.08mg/L) hypoxia. Hypoxia acted similar to an estrogen mimic, with huge up-regulation of VTG gene expression in males, and increased VTG levels (additive effect) when hypoxia was combined with 4tOP. Microarray analyses showed 125 genes with significant transcriptional change, with up- or down-regulation from transitions of: (1) hypoxia (72 hrs) to normoxia (74 hrs) and (2) hypoxia+4tOP (72 hrs) to normoxia+4tOP (74 hrs).