The 1,2,5-Thiadiazolidin-3-one 1,1-dioxide scaffold was used in the design and synthesis of inhibitors of Dengue Virus and West Nile Virus proteases and human tryptase. The scaffold was successfully used in the synthesis of potential inhibitors of Dengue Virus and West Nile Virus proteases. Inhibitors of Human tryptase synthesized based on the 1,2,5-Thiadiazolidin-3-one 1,1- dioxide scaffold were shown to be effective mechanism-based inhibitors of the enzyme. / Thesis (M.S.)--Wichita State University, College of Liberal Arts and Sciences, Dept. of Chemistry. / "July 2006." / Includes bibliographic references (leaves 64-66).

The groupers are vital fish in Taiwan, the market of grouper fry over 300 million dollars. While grouper nervous necrosis virus (NNV) has caused mass mortality, especially 100% in larvae and juveniles, which economically impacts on culture of marine fish. The vaccination is one of the best methods to against viral diseases. The dragon grouper (Epinephelus lanceolatus), malabar grouper (E. malabaricus) and brown-marbled grouper (E. fuscoguttatus) were injected with different dosages and injection frequencies of virus-like particles (VLPs) of DGNNV, which is by the first claimed. The anti-sera of vaccinated fish were analyzed with eight kinds of immunology methods, among which antigen-capture ELISA was the best choice for qualitative and quantitative assays. The signal of antibodies in the vaccinated fish was detected in all groupers in one week after primary immunization, and the antibody titers increased markedly in one month. In dragon grouper, fish was injected with 10 £gg of VLPs, the antibody titer reached 1.05. To given booster injection once, antibody titers were raised to 35.7%. In malabar grouper, after injected twice with 50 £gg of VLPs, the antibody titer raised 33.3% than inoculation once in six weeks. After brown-marbled grouper was injected with 450 £gg of VLPs, the high antibody titer reached to 1.57 at five weeks post primary immunization. Specific antibodies still can be detected after seven months. In the in vitro assay with MGNNV of 103.5 TCID50/mL, neutralizing antibody titer of control fish were all lower than 1:50. The neutralizing antibody titer of anti-serum of dragon grouper was detected at 1:200 at one week, and raised to 1:1600 at four weeks and 1:6400 at eleven weeks after primary vaccination. In malabar grouper and brown-marbled grouper, the neutralizing antibody titers were 1:3200 and 1:400, respectively, in one month. The antibody titer can not increased by Freund¡¦s complete adjuvant. The fish produced high antibody titer and high protection by immunization with VLPs.

El objetivo del presente estudio fue evaluar el grado de susceptibilidad, el efecto patológico y respuesta serológica de la paloma silvestre (especie Columba livia) frente a la inoculación experimental con una cepa de virus velogénico de la enfermedad de Newcastle. Se capturaron 28 aves adultas, 14 fueron inoculadas con una cepa velogénica viscerotrópica de la enfermedad de Newcastle, vía nasal y oral, mientras las restantes fueron criadas como grupo control. En las aves inoculadas se registró los signos clínicos y mortalidad. Se tomaron muestras de diferentes tejidos de aves que murieron así como del grupo control, para su análisis histopatológico. Se tomaron muestras de pulmón y tráquea e hisopado de cloaca para la recuperación viral, semanalmente durante seis semanas. El 64% de aves del grupo inoculado presentó signos clínicos y se obtuvo una mortalidad del 42.8%. Se presentaron estornudos a partir del cuarto día post inoculación. El erizamiento de plumas, aislamiento y letargia, se observó a partir del quinto día en aves que presentaron signos respiratorios y nerviosos. Al séptimo día se observó en el 42.8% opistótonos, tremores de cabeza y cuello que se exacerbaban al estímulo de ruidos. Al octavo día sólo en el 21 % de las aves mostraron diarrea. Los hallazgos a la necropsia consistieron en una congestión generalizada de los diferentes órganos y esplenomegalia. Las lesiones microscópicas más marcadas fueron edema, gliosis, manguito perivascular a mononucleares en cerebro y cerebelo, pérdida de cilios, infiltrado de linfocitos en tráquea, congestión interalveolar en pulmón, congestión en proventrículo, congestión e infiltración de linfocitos en intestinos y despoblamiento linfoide en bazo. El grupo inoculado mostró un incremento en sus títulos de anticuerpos a partir de la primera semana post inoculación llegando a su máximo nivel promedio geométrico de título (P.G.T) de 4.9 en la segunda semana para luego descender por completo a la quinta semana. Se logró la recuperación viral a partir de muestras de pulmón y tráquea durante las tres primeras semanas post inoculación. Se demostró que la especie Columba livia es susceptible a la inoculación experimental con una cepa velogénica del virus de la enfermedad de Newcastle. / This study was designed to asses the susceptibility, pathologic effect and serologic response of wild pigeons (sp. Columba livia) to Newcastle virus. A total of twenty eight adult wild pigeons were captured, fourteen were inoculated with a velogenic viscerotropic strain of Newcastle virus by oral and nasal route. The remaining pigeons were used as control group. The birds were observed to record clinical signs and mortality, also blood samples were collected for hemaglutination inhibition technique and tissue samples from lungs, tracheas and cloacal swabs were harvested for the viral recovery and histological studies. The 64 % of the inoculated group showed clinical signs and a mortality of 42.8 %. The clinical signs (sneezes, ruffled plumage, isolation and lethargy) started the fourth day post inoculation. The 43 % showed nervous signs: opisthotonos, tremors of head and neck that was exaggerated to the stimulus of noises and the 21 % showed diarrhea. In the necropsy were observed a widespread congestion and splenomegaly. The microscopic injuries were edema, gliosis, mononuclear perivascular cuffing in brain and cerebellum, loss ciliar, lymphoid infiltration in trachea, lungs congestion, proventricular congestion, and lymphocitic infiltration in intestines and lymphoid depletion in spleen. The inoculated group showed an increase in the antibody titers the first week post inoculation reaching the highest titer mean (P.G.T=4.9) in the second week after this they descended. The viral recovery was made upon lungs and tracheas tissues and from a cloacal swab in dead birds. It was showed that the specie Columba livia was susceptible to the experimental inoculation with a velogenic strain of Newcastle diasease virus.

El presente trabajo tuvo como objetivo estandarizar y validar la técnica de RT-PCR en tiempo real para el diagnóstico del virus de la Necrosis Pancreática Infecciosa (IPNV) en la trucha arcoíris (Oncorhynchus mykiss) en el Perú. Se utilizaron muestras de riñón y bazo de truchas arcoíris provenientes de dos piscigranjas del departamento de Junín, tomándose 61 animales con signos clínicos de enfermedad y 60 animales aparentemente sanos. Todas las muestras fueron evaluadas mediante la técnica de Inmunofluorescencia Indirecta (IFI) para determinar la presencia del virus. La prueba de RT-PCR tiempo real se realizó utilizando un kit comercial en dos pasos y utilizando el fluoróforo Sybr Green I. Se utilizaron los primers WB1 y WB2 para identificar un segmento genómico específico de la proteína estructural VP2. Como controles positivos se utilizaron virus inactivado IPNV cepa Sp, así como 10 muestras inoculadas con IPNV; y como controles negativos, se utilizaron virus relacionados a IPNV como son Rotavirus A y el virus de la Bursitis Infecciosa del pollo; así como virus no relacionados. A su vez, se llevó a cabo la inoculación viral en muestras de tejido en diluciones decrecientes para determinar el grado de sensibilidad. Los resultados fueron determinados a partir de los valores del Ciclo Umbral (Ct) y temperatura de Melting (Tm) de los productos amplificados. La prueba fue capaz de detectar al virus hasta concentraciones de 102 UFP/ml. Las 121 muestras de campo resultaron negativas a IPNV; los animales enfermos evidenciaron etiologías de tipos bacterianos y no virales. Los resultados obtenidos determinaron una especificidad del 100% y una sensibilidad del 100%; a su vez, los valores predictivo positivo y predictivo negativo resultaron en 100%. La técnica de RT-PCR en tiempo real estandarizada en el laboratorio representa una técnica valiosa para el diagnóstico de IPNV. Palabras Clave: Virus de la Necrosis Pancreática Infecciosa, IPNV, RT-PCR, RT-PCR tiempo real / This study aimed to standardize and validate the real-time RT-PCR technique for diagnosis of Infectious Pancreatic Necrosis Virus (IPNV) in rainbow trout (Oncorhynchus mykiss) in Peru. Kidney and spleen samples of rainbow trout (n=121) from two fish farms of Junin were taken, 61 animals showed signs of disease and 60 were apparently healthy animals. All samples were assessed by indirect immunofluorescence technique (IIF) to determine presence of virus. Real-time RT-PCR was carried out using a two-step commercial kit. We used WB1 and WB2 primers for amplification of a specific genomic segment of the VP2 structural protein. IPNV Sp strain and 10 samples inoculated with IPNV were used as positive controls; whereas related viruses, such as rotavirus and infectious bursal disease virus from chickens, as well as unrelated viruses were all used as negative controls. Also, we inoculated decreasing dilutions of virus into tissue samples to determine the sensitivity degree of the test. Results were assesssed according to the values of the threshold cycle (Ct) and melting temperature (Tm) of the amplified products. The test was able to detect the virus in concentrations up to 102 PFU/ml. All 121 field samples were negative for IPNV; sick animals were apparently infected by bacterial agents. Real Time RT-PCR’s specificity and sensitivity were around 100%; moreover, the positive predictive value and negative predictive value was also set in 100%. Real time RT- PCR technique standardized in our laboratory is a valuable diagnostic tool for IPNV detection in field samples. Key Words: Infectious Pancreatic Necrosis Virus, IPNV, RT-PCR, real time RT-PCR

Thèse de doctorat : Biologie moléculaire : Strasbourg 1 : 2007. / Thèse soutenue sur un ensemble de travaux. Titre provenant de l'écran-titre. Bibliogr. f. 116-131.

Dissertation (Ph.D.)--Auburn University, 2009. / Abstract. Includes bibliographic references (p.72-81).

Title from PDF of title page (University of Missouri--Columbia, viewed on Feb 19, 2010). The entire thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file; a non-technical public abstract appears in the public.pdf file. Thesis advisor: Dr. James E. Schoelz. Includes bibliographical references.

Thesis (doctoral)--Universität, Münster (Westfalen).

Thesis (Ph.D.)--Memorial University of Newfoundland, 2002. / Bibliography: leaves 221-251.

published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy