Bacterial polyketides possess an enormous range of chemical diversity and biological function. Many polyketides such as tetracycline, epothilone, and rapamycin have been developed into key clinical pharmaceuticals in a broad range of therapeutic areas. Sequencing of bacterial genomes has shown that there are many more polyketide biosynthetic pathways than there are polyketides isolated from standard cultivation techniques. These genetically encoded polyketide natural products from cultivatable and uncultivatable bacteria represent one of the greatest remaining untapped reservoirs of new natural product diversity. To access this untapped diversity of polyketide products, a general method for heterologous expression of these pathways is needed. Heterologous expression has proven to be a valuable asset in the discovery, production, engineering, and characterization of bacterial secondary metabolites and the complex enzymology involved in their biosynthesis. Herein we discuss the development and investigation of two unique heterologous expression platforms utilizing host strains of Myxococcus xanthus and Escherichia coli. Using our developed heterologous hosts, we were able to produce the Streptomyces rimosus polyketide oxytetracycline. Through production of oxytetracycline in E .coli we have identified the potential of alternative transcription factors as regulators of secondary metabolism. Further investigation and development of alternative transcription factors as regulators of secondary metabolism in heterologous hosts could benefit the development of robust general methodology for the heterologous expression of polyketides.

Mississippi is home to many small impoundments, providing anglers with local fishing opportunities. Crappie (Pomoxis spp.) fisheries are highly prized in the state, and managers struggle to meet angler expectations in these smaller systems. From 2015 to 2017, white crappie (Pomoxis annularis) and magnolia crappie (male black crappie Pomoxis nigromaculatus x female white crappie Pomoxis annularis), marked with oxytetracycline and alizarin red s, were stocked into five small Mississippi impoundments to evaluate stocking efficacy and long-term mark retention. Overall stocking contribution to established populations was 0.59% with only one marked fish recovered. An additional 12 lakes ranging in size from 28 and 357 hectares were studied to assess factors contributing to successful crappie populations. Highest crappie growth rates were observed in systems where crappie abundance is limited. Findings of this study suggest predator population management can be a valuable tool for managing crappie in small impoundments.

Summer flounder, Paralichthys dentatus, culture is becoming increasingly popular in the United States because of high market prices and consumer demand. In addition, flounder is a marine fish species that can tolerate a wide range of salinities, allowing for inland intensive fish culture. Oxytetracycline (OTC) is one of two available FDA-approved antibiotics for use in foodfish in the United States. Oxytetracycline was chosen for these studies because it is excreted primarily unchanged through the urine and the absorption, distribution and elimination of this drug may be influenced by environmental and physiological conditions. Four experiments were conducted to investigate: 1) pharmacokinetic parameters of oxytetracycline (50 mg/kg) following intravascular (IV), intraperitoneal (IP), intramuscular (IM) and per os (PO) administration in summer flounder maintained at 28 ppt salinity and 20°C; 2) pharmacokinetic parameters of OTC (50 mg/kg) following IM and PO administration in summer flounder maintained at three different salinity levels of 0 ppt, 15 ppt and 32 ppt and the physiological adjustments summer flounder make to acclimate to environmental salinity; 3) OTC retention times in muscle tissue from summer flounder maintained at three different salinity levels (0 ppt, 15 ppt, 32 ppt) and treated with a single 50 mg/kg OTC dose via IM and PO administration; and 4) pharmacokinetic parameters of OTC (50 mg/kg) following IM and PO administration in clinically healthy and clinically diseased summer flounder maintained at 28 ppt and 20°C. Oxytetracycline plasma concentrations were determined using high performance liquid chromatography (HPLC) and analyzed using a non-compartmental pharmacokinetic model for all routes of drug administration. Statistical comparisons were not made between the different routes of OTC exposure, but results from experiment one indicated that IV administration of OTC resulted in the largest area under the curve (AUC) value (8147.9 µg·h/ml) and the highest maximum plasma concentration (Cmax) of 1173.2 µg/ml OTC at 5 min post-injection. Intramuscular injections of OTC resulted in prolonged total body elimination half-life (T ½) of 301.3 h and high fish-to-fish variability (0.6). Per os administration resulted in low Cmax (0.2 µg/ml OTC) and poor systemic bioavailability (0.2 %). Results from experiment two demonstrated that when OTC is administered IM AUC estimates are significantly (p<0.05) lower in summer flounder held at 0 ppt (1684.8 µg·h/ml) than fish maintained at 15 ppt or 32 ppt salinity (2067.8 µg·h/ml and 2241.3 µg·h/ml, respectively). Although not significantly different from other salinity treatments, time to maximum plasma concentration (Tmax) was longer in fish held at 15 ppt and 32 ppt (312 h and 168 h, respectively) compared to cohorts in freshwater (0.5 h) and Cmax values were higher in animals held at 15 ppt and 32 ppt (8.4 µg/ml OTC and 9.2 µg/ml OTC, respectively) than freshwater fish (4.9 µg/ml OTC) when OTC was administered via IM injection. No significant differences were detected in any of the pharmacokinetic parameters following PO dosing of OTC, however, the AUC estimates were lower in the 32 ppt acclimated fish (127.7 µg·h/ml) than in the 0 ppt or 15 ppt acclimated fish (190.2 µg·h/ml and 180.7 µg·h/ml, respectively). In addition, the T ½ was longer in the higher salinity groups (278.1 h and 266.0 h, respectively) than in the freshwater fish group (256.9 h). Physiological adjustments made by summer flounder including plasma and urine osmolality, urine flow rate and urine character, gill chloride cell size and density, and Na+ - K+ ATPase activity demonstrated trends that suggested physiological differences among the salinity groups. Plasma and urine osmolalities were typically significantly (p<0.05) higher in fish maintained at 32 ppt salinity than at the lower salinity treatments. In addition, urine flow rates were generally significantly (p<0.05) greater in freshwater adapted fish (0.13 - 0.21 ml of urine/kg/hour) in comparison to fish in the salinity treatments of 15 ppt and 32 ppt (0.06 - 0.12 ml of urine/kg/hour and 0.09 – 0.11 ml of urine/kg/hour, respectively). Changes in gill chloride cell size and density and enzyme activity of Na+ - K+ ATPase revealed no significant differences between the salinity treatments but summer flounder in saltwater had numerically larger and more chloride cells than summer flounder in freshwater, but enzyme activity was greater in freshwater acclimated summer flounder compared to fish in seawater. Experiment three results revealed similar OTC muscle tissue pharmacokinetic parameters in summer flounder following IM injection. However, there were significant differences (p<0.05) in the AUC parameters of the plasma and muscle OTC concentrations between fish maintained at different salinities following IM OTC treatment. These effects may be the result of a "depot" effect in the muscle tissue or may be related to the reduced solubility of OTC in the muscle tissue of marine fish. A single PO dose administration of OTC at 50 mg/kg did not result in plasma or tissue concentrations higher than the FDA tissue tolerance limit of 2 ppm. Results of the fourth experiment demonstrated that following IM OTC administration healthy fish had significantly (p<0.05) higher AUC (4700.6 µg·h/ml) values than diseased cohorts (2576.2 µg·h/ml). Maximum plasma concentrations were also higher in the healthy fish than in the diseased fish, although values were not significantly different (23.4 µg OTC/ml and 20.2 µg OTC/ml, respectively for healthy and diseased fish). Additionally, in diseased fish, the mean resident time (MRT) (293.7 h) and T ½ (203.5 h) parameters were longer compared to parameters in healthy fish (253.2 h and 175.4 h, respectively), although values were not significantly different. No significant differences were detected in any of the pharmacokinetic parameters following PO OTC administration, however, healthy fish achieved higher maximum plasma OTC concentrations (1.0 µg OTC/ml) than diseased fish (0.7 µg OTC/ml). Fish-to-fish variation was greater in diseased animals than in healthy regardless of route of drug administration. The results of these experiments indicated that OTC pharmacokinetic parameters are influenced by route of drug administration, environmental salinity and fish health status. These factors must be considered by veterinarians and governmental regulators when developing treatment regimens for summer flounder. / Ph. D.

No description available.

Graduation date: 1999

No description available.

The specificity of a medaka vitellogenin (Vtg) enzyme-linked immunosorbent assay (ELISA) kit was tested on adult hybrid tilapia to determine if vitellogenin could be successfully measured in tilapia using this method. Adult hybrid (Orechromis niloticus x Oreochromis mossambicus x Oreochromis aureus) tilapia were injected with 17â-estradiol (E2) at 5 ìg/g body weight (b.w.) in two separate experiments and exposed to 2.5 ìg/g b.w. E2 through aquaria water in a third experiment. All fish were exposed to solvent controls (50/50 PBS/ethanol, or 59/41 PBS/ ethanol) proportional to the volume of estradiol solvent used. Experiment 1 was terminated due to a decrease in fish health and unexpected mortalities. Experiment 2 was continued though there was again a decrease in fish health and mortalities before experiment was completed. According to one-way ANOVA, Experiment 2 treatment groups were significantly different (P<0.05); however, Tukey's pairwise comparisons showed no differences between treatment groups. Experiment 3 results showed statistical differences between some treatment groups (P<0.001). However, maximum vitellogenin levels reached 1250 ng/mL Vtg, which were much lower than induced vitellogenin levels from literature (0.002 mg/mL to 6 mg/mL). Typical induction trends should be as follows: male control<female control< induced male<induced female; however, these trends were not observed in this study. For these reasons, the medaka vitellogenin ELISA kit was determined not specific to tilapia vitellogenin. This research was performed due to the fact that tilapia are farmed worldwide, and to increase research with these fish, in order to better understand their response to xenobiotic chemicals, easier methods are desired for measuring their vitellogenin. Vitellogenin induction from exposure to the chemotherapeutics: oxytetracycline, Romet-30®, and copper sulfate was evaluate using Japanese medaka (Oryzias latipes). Medaka were exposed to E2 (0.0552 ìg/L) in aquaria water as a positive control. Medaka were fed oxytetracycline medicated feed at 0.0174 g, 0.0348 g, and 0.087 g, and Romet-30® medicated feed at 0.0261 g, 0.0522 g, and 0.1305 g daily for 11 days and 5 days as recommended. Medaka were exposed to copper sulfate at 1.85 mg/L for one hour (for 1 day, 7 consecutive days, and every three to five days for a total of 7 doses). Vitellogenin levels were measured using a sandwich ELISA. As expected, Vtg levels were induced from exposure to E2 (P<0.01); however, exposure to the chemotherapeutic compounds did not induce elevated Vtg levels. Alternately, copper sulfate exposure for one hour for 1 day did show a significant decrease in vitellogenin (P<0.01). Biological variation accounted for large portions of the variation within treatment groups in medaka vitellogenin in fish exposed to chemotherapeutics (as high as 41.01%). From this research we suggest that these chemicals do not have estrogenic properties capable of inducing vitellogenin in medaka. / Master of Science

Thesis (M.S.)--Oregon State University, 2007. / Printout. Includes bibliographical references (leaves 35-40). Also available on the World Wide Web.

Thesis (M.Sc.), Memorial University of Newfoundland, 2000. / Bibliography: leaves 69-72.

Orientador: Felix Guillermo Reyes Reyes / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-09T00:18:19Z (GMT). No. of bitstreams: 1 Maia_PatriciaPenido_D.pdf: 738805 bytes, checksum: 45cdc642b12e037677eca58fb443a574 (MD5) Previous issue date: 2007 / Resumo: Antimicrobianos como fungicidas e/ou bactericidas têm sido utilizados para a melhoria da qualidade de produtos vegetais destinados à alimentação humana ou para sua maior produtividade em várias fases de sua obtenção, tais como: preparo do solo, crescimento, colheita, industrialização e comercialização. Entretanto, vários destes compostos podem permanecer como resíduos nos alimentos consumidos, e, dependendo de sua toxicidade e nível de exposição, podem apresentar risco à saúde do consumidor. Entre estas substâncias estão os agentes antimicrobianos usados para combater ou prevenir o aparecimento de alterações de origem microbiana no alimento. O desenvolvimento de métodos analíticos para a identificação e quantificação destes agentes em produtos vegetais é de grande interesse, uma vez que a exposição crônica em baixos níveis antimicrobianos pode promover a resistência bacteriana a antibióticos, assim como reações alérgicas. Dessa forma, o presente trabalho objetivou desenvolver metodologia para o isolamento, identificação e quantificação de resíduos de oxitetraciclina em tomate, o qual é muito sensível a bacterioses, assim como comparou os níveis residuais encontrados nos frutos de cultura protegida (estufa) e cultura de campo. A metodologia analítica desenvolvida foi aplicada na análise de tomates comercializados na região de Campinas/SP e Alfenas/MG. Os dados obtidos foram comparados com os limites máximos de resíduos (LMRs) e os intervalos de segurança estabelecidos pela legislação brasileira em condições de plantio no campo. Foi validado, ainda, um método para quantificação dos antibióticos oxitetraciclina e estreptomicina em formulações para uso agrícola utilizadas para tratamento de tomates / Abstract: Antimicrobials as fungicides and/or bactericides have been used for the improvement of the quality of vegetal products destined to the human feed and for its higher productivity in some phases of its cultivation, such as: preparation of the ground, growth, harvest, industrialization and commercialization. However, several of these compounds can remain as residues in consumed foods, and, depending on its toxicity and level of exposure, they can present a risk to the health of the consumer. Among these substances are the antibiotics used to combat or to prevent the appearance of alterations of microbial origin in the food. The development of analytical methods for the identification and quantification of these agents in vegetal products is of great interest, once the chronic exposure to low levels of antimicrobials can promote bacterial resistance to the antibiotics, as well as allergic reactions. Therefore, the present study aimed to develop a methodology for the isolation, identification and quantification of residues of oxytetracycline in tomato, which is very sensible to bacterioses, as well as was compared the residual levels found in fruits cultivated in greenhouse and open field. The developed analytical methodology was applied in the analysis of tomatoes purchased in the region of Campinas/SP and Alfenas/MG. The obtained data were compared with the maximum residue limits (MRLs) and the intervals of security established by the Brazilian legislation in conditions in the field. It was validated a method for quantification of antibiotics oxytetracycline and streptomycin in agricultural formulations used for treatment of tomatoes / Doutorado / Doutor em Ciência de Alimentos