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Purpose: Translational research characterizing endothelial dysfunction and the progression of cardiovascular disease (CVD) is necessary for understanding the complex nature of multi-factorial diseases. Perhaps more important though, is understanding the compensatory and adaptive processes associated with regression of diseases and chronic oxidative stress. Vascular endothelial growth factor (VEGF) is an important protein in endothelial health and nitric oxide (NO) production. The purpose of this research was to examine changes in VEGF-mediated endothelial nitric oxide synthase (eNOS) activity under conditions of oxidative stress both in vivo and in vitro. Methods: The oxidative stress relationship involving plasma VEGF, NO, and hydrogen peroxide (H2O2) was assessed in sedentary, pre-hypertensive African American participants both (n=48) before and following (n=22) 6 months of aerobic exercise training (AEXT). In vitro, H2O2 exposure along with atherogenic, 4 dyne/cm2, and athero-protective, 20 dyne/cm2, levels of laminar shear stress (LSS) were used to characterize VEGF-mediated eNOS activity to gain insights into physiological signaling. Results: At baseline, VEGF levels increased with increasing blood pressure (BP) level while NO levels decreased from normotensive to hypertensive participants. H2O2 levels also trended upward with increasing BP level, and in vitro H2O2 was observed to decrease VEGF-mediated eNOS activity in a dose dependent manner. Following AEXT, participants were divided into groups relative to their BP change following the intervention. Participants that decreased their BP level demonstrated a decrease in VEGF and H2O2 level. In addition, following 24 hrs of LSS at 20 dyne/cm2, VEGF-mediated eNOS activity and VEGFR2 protein expression was significantly lower compared to 24 hrs of LSS at 4 dyne/cm2. Discussion: Increased circulating levels of VEGF in vivo may be a compensatory mechanism. Endothelial dysfunction and progressive CVD may trigger such compensation. The adaptive response to exercise for its BP-lowering effects is systemic and encompasses many changes. These beneficial adaptations have likely alleviated the compensatory mechanism of elevated VEGF levels seen at baseline. Indeed, following 24 hrs of an athero-protective LSS level, VEGF-mediated eNOS activity was significantly lower compared to 24 hr of LSS at an atherogenic level. The difference in VEGF-mediated eNOS activity may be due, in part, to the decrease in VEGFR2 protein expression we observed under an athero-protective LSS level. / Kinesiology

Chill brines used in ready-to-eat meat processing may be an important source of post-processing contamination by Listeria monocytogenes. The purpose of this study was to determine the efficacy of ultraviolet light (UV) in combination with antimicrobials to reduce L. monocytogenes in fresh and used chill brines. Three different antimicrobials were used in combination with UV; citric acid (CA, 0.2 and 0.5%), dimethyl dicarbonate (DMDC, 250 and 500 ppm), and hydrogen peroxide (HP, 2000 and 4000 ppm). For fresh brine studies, brine (8.0% w/v NaCl) was prepared and inoculated with a cocktail of three L. monocytogenes strains (approximately 6 log CFU/mL). Brine was treated with UV alone, antimicrobials alone, and combination of UV and antimicrobials. Moreover, to observe the effect of treatment temperature and brine circulation through the UV system on survival of listeriae cells, inoculated brine was circulated through the system without any treatment that served as control for all the treatments. For UV treatment, inoculated brine solution was exposed to UV in an Ultraviolet Water Treatment Unit (Model: AMD 150B/1/2T D; Aquionics Inc., Peak output: 254 nm) fitted with an inline chiller to maintain brine temperature of -1°C. Samples were withdrawn at regular intervals for 120 minutes. When L. monocytogenes population was no longer detectable via direct plating on MOX, enrichment was performed and suspect colonies were confirmed using API-Listeria. For antimicrobial-only (i.e., no UV) treatments, a specific concentration of antimicrobial was added in inoculated brine and samples were taken for 120 minutes. For the brine that received combination of UV and antimicrobial treatments, UV was turned on once a specific concentration of antimicrobial was added in inoculated brine and samples were withdrawn at regular intervals for 120 minutes. When treated with UV alone, L. monocytogenes population decreased from approximately 6 log CFU/mL to below the detection limit (i.e., 1 log CFU/mL) in 15 minutes with the reduction rate of 0.87 log CFU/mL per minute. However, cells were detectable by enrichment through 120 minutes. The highest rate of decline (0.90 log CFU/mL per minute) was achieved by the combination of UV and 500 ppm DMDC (UV+500 ppm DMDC), which was not significantly different from the reduction rates of UV and UV+0.5% CA. UV+500 ppm DMDC reduced L. monocytogenes to the detection limit in 15 minutes and the organism was not detected by enrichment after 60 minutes. Though the reduction rate of UV+0.5% CA was not significantly lower than the rate of UV+500 ppm DMDC (P>0.05), the former treatment resulted in non-detectable levels more quickly (45 minutes) than the latter (60 minutes). Thus, based on enrichment studies UV+0.5% CA was the most effective treatment in reducing the population of L. monocytogenes in fresh brine. Moreover, when brine was treated with 0.5% CA alone the population decreased to below detection limit in 15 minutes with the rate significantly lower than UV+500 ppm DMDC and UV+0.5% CA (P<0.05). However, L. monocytogenes was not detectable by enrichment from 60 minutes. To summarize, through enrichment studies we observed that UV+0.5% CA, UV+500 DMDC, and 0.5% CA Control were more effective than other treatments in reducing the listeriae population to a non-detectable level. Spent brine is recycled brine that was obtained from a frankfurter processor after its maximum usage. Results of spent brine studies showed that when brine was treated with UV+4000 ppm HP and UV+2000 ppm HP, L. monocytogenes population decreased to the detection limit in 45 minutes and was not detected by enrichment from 120 minutes. These treatments were observed to be the most effective treatments with a reduction rate of 0.12 log CFU/mL per minute. The reduction rate of some other treatments such as, UV+250 and 500 ppm DMDC, UV+0.2% and 0.5% CA, and UV alone was not significantly different from UV+4000 and 2000 ppm HP. However, the population was detected through enrichment up to 120 minutes in all other treatments. The results of these studies indicate that combinations of UV and antimicrobial may be more effective than either treatment alone (except 0.5% CA treatment) to process fresh chill brines. However, the antimicrobials and UV were less effective for controlling L. monocytgoenes in spent brine; presumably due to the presence of organic matter. / Ph. D.

Hazard Analysis Critical Control Point (HACCP) regulations for juice manufacture require the application of a process that will result in a 5-log reduction (99.999%) of the pertinent pathogen in the juice being processed. The use of ultraviolet (UV) light, as an alternative to traditional thermal processing, has been adopted by some juice processors as a means of meeting the HACCP 5-log performance standard. However, little research had been performed to determine the effect of UV when used in combination with antimicrobial agents that are commonly added to juice products. Therefore, the objectives of this work were (1) to determine if chemical preservatives and ultraviolet light have a combined effect on the reduction of Escherichia coli in apple cider, and (2) to determine the influence of adding chemical preservatives at different points in the processing of juice (i.e., either prior to or after ultraviolet light processing) on the reduction of Escherichia coli in apple cider. In this study, refrigerated (4°C) pasteurized apple cider that contained no added preservatives was inoculated with E. coli ATCC 25922, a surrogate strain for E. coli O157:H7, and exposed to UV (peak output: 254 nm). The following chemical preservatives were added to apple cider either prior to or after UV exposure: dimethyl dicarbonate (75 and 150 ppm), hydrogen peroxide (75 and 150 ppm), potassium sorbate (1000 and 2000 ppm), and sodium benzoate (1000 and 2000 ppm). Following UV exposure and chemical preservative application, inoculated juices were stored at 4°C for 72 hours. Samples were collected prior to and immediately after UV exposure and at 24, 48, and 72 hours of storage. At each sampling point, juice portions (0.1 ml) were serially diluted in peptone diluent (0.1%) and surface plated onto Tryptic Soy Agar (TSA). Counts of the bacterial colonies were made 48 hours after incubating plates at 35°C. Overall, reductions of E. coli were greater in cider treated with preservatives after UV processing than when preservatives were added prior to UV processing (P < 0.05). Furthermore, dimethyl dicarbonate and hydrogen peroxide were more effective than potassium sorbate and sodium benzoate in reducing E. coli populations in conjunction with UV (P < 0.05). When added prior to UV exposure, potassium sorbate was the least effective, allowing for the greatest survival (P < 0.05). This study describes the use of UV in conjunction with hydrogen peroxide and dimethyl dicarbonate as an effective method for producing a 5-log or greater reduction of E. coli O157:H7 in apple cider. / Master of Science

The pathogenic microorganism Salmonella enterica has been associated with several outbreaks and recalls of spices, herbs, and seeds. To control these pathogens additional treatment methods, such as fumigation with chlorine dioxide (ClO2) or hydrogen peroxide (H2O2) gas and recovery methods are needed. Recovery methods should accurately quantify all viable cells, even those injured, to prevent overestimation of treatment effectiveness. This study was performed to determine the effect of different recovery media and supplements on the recovery of multiple strains of S. enterica and Enterococcus faecium NRRL B2354, from chlorine dioxide or hydrogen peroxide treated low moisture foods (LMF) black peppercorns, dried basil leaves, and chia seeds. Also, this study aimed to compare the log reduction of these two microorganisms to evaluate E. faecium NRRL B2354 as a surrogate for S. enterica. On average, recovery of S. enterica was 3.43 log and 4.77 log CFU/g from ClO2 and H2O2 treated LMFs, respectively on the selective media Xylose Lysine Deoxycholate agar, while the average recovery on non-selective media was 4.50 log CFU/g and 5.74 log CFU/g from ClO2 and H2O2 treated LMFs, respectively. The use of non-selective media was correlated with increased recovery compared to selective media. In further studies, addition of sodium pyruvate, ferrous sulfate, or 3'3'-thiodiproionate supplements to MTSAYE did not show increased recovery (P>0.05). On each treatment and LMF combination tested, there was no significant difference between the log reduction of S. enterica and E. faecium NRRL B2354, indicating its suitability as a surrogate under the test conditions. / Master of Science in Life Sciences / Spices, dried herbs, and seeds have become popular throughout the world for enhancing the flavor of food, but may also harbor harmful bacteria, including Salmonella enterica. It is US federal law under the Food Safety Modernization Act that these foods are safe to eat straight from processors since these foods are typically consumed raw. Novel treatment methods are being tested to kill harmful bacteria on these dried foods without adding water including chlorine dioxide fumigation and hydrogen peroxide fumigation. However, these processes can injure the bacteria without killing them. These injured bacteria might not be counted using traditional means which could lead to overestimating the effectiveness of a treatment. Different media types, used as part of the process to count the number of bacteria in a sample, were tested to determine their effect on recovery of injured S. enterica cells. Furthermore, the bacterium Enterococcus faecium NRRL B2354 was tested against S. enterica to evaluate, if the former, a relatively harmless microorganism, could be used by food processing plants to determine that their treatment processes meets regulatory standards. More injured S. enterica cells were recovered from each non-selective media tested, compared to the selective media. Although there isn't a significant difference in injured S. enterica recovery between any supplemented non-selective media, any non-selective media recovers more sub-lethally injured cells, and would give more accurate bacterial counts. Results also indicated that E. faecium NRRL B2354 is a suitable surrogate to the pathogen S. enterica for spices and herbs processed under the same conditions.