Thesis (Ph. D.)--University of Wisconsin--Madison, 1963. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves [77]-85).

Made available in DSpace on 2014-06-11T19:27:16Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-06-30Bitstream added on 2014-06-13T20:35:29Z : No. of bitstreams: 1 fuverki_rbn_me_jabo.pdf: 380911 bytes, checksum: 1e3cde0d1d852b172186ddec592ea827 (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Com a crescente disponibilidade das biotécnicas de reprodução animal, o comércio dos produtos envolvidos com essas práticas também está em expansão, oferecendo a possibilidade de melhoria dos índices zootécnicos às produções de bovinos. Porém deve-se levar em consideração que há riscos sanitários em práticas como a inseminação artificial caso não se realize o controle do material biológico utilizado. Dentre os agentes infecciosos que podem estar presentes no sêmen e passíveis de serem transmitidos por esse estão a leptospirose e a brucelose, enfermidades responsáveis por grandes perdas reprodutivas e econômicas na bovinocultura mundial. Este projeto teve como objetivos detectar molecularmente esses patógenos em amostras de sêmen bovino provenientes de centrais de comercialização brasileiras, utilizando um kit comercial para extração de DNA (“RTP Bacteria DNA Mini Kit” (Invitek®), aperfeiçoá-lo para a extração de DNA bacteriano a partir de sêmen e avaliar sua aplicabilidade à rotina laboratorial. O DNA bacteriano foi extraído e quantificado por eletroforese em gel de agarose. Pretendeu-se também realizar reação em cadeia da polimerase (PCR) utilizando os “primers” B4 e B5 para amplificação do DNA de Brucella spp. e os “primers” Lep 1 e Lep 2 para Leptospira spp. O kit de extração foi otimizado com sucesso, e todas as 96 amostras examinadas foram negativas para qualquer DNA bacteriano. Os resultados podem ser úteis para estabelecer alternativas de controle sanitário em touros doadores de sêmen e permitir o fornecimento de material genético livre de patógenos, aumentando o “status” sanitário da reprodução de bovinos no Brasil / With the increasing disponibility of animal reproduction biotechniques, trading of products involved with these activities is also in expansion offering improving possibilities in zootecnic indexes of bovine herds. However, considerations should be taken about sanitary risks in practices like artificial insemination if any control is applied to this biological material. Among infectious agents that could be present and transmitted by semen are leptospirosis and brucellosis, diseases that are responsible for numerous reproductive and economic losses in world’s cattle culture. The goals of this project were to molecularly detect these pathogens in bovine semen samples from Brazilian artificial insemination centers using a commercial kit for DNA extraction (RTP Bacteria DNA Mini Kit (Invitek®), to improve it for extracting bacterial DNA from semen and to analyze its applicability in laboratory routine. Bacterial DNA was extracted and quantified by agarose gel electrophoresis. We also intended to realize polymerase chain reaction (PCR) using primers B4 and B5 for amplification of Brucella spp. DNA and primers Lep 1 e Lep 2 for Leptospira spp. DNA. The extraction kit was successfully optimized and all of 96 examined samples were negative for any bacterial DNA. Results could be useful to establish alternative measures of sanitary control in semen donors and to allow the supplying of genetic material free of pathogens, increasing sanitary status of bovine reproduction in Brazil

Memoria para optar al Título Profesional de Médico Veterinario / La Brucelosis canina (BC) es una enfermedad infectocontagiosa zoonótica causada principalmente por la bacteria Brucella canis (B. canis). Sin embargo, se han descrito infecciones esporádicas causadas tanto por Brucella suis (B. suis) y Brucella abortus (B. abortus) como también por Brucella melitensis (B. melitensis), las cuales producen en el animal un cuadro autolimitado. La infección se caracteriza por producir infertilidad tanto en machos como en hembras, afectando la vida reproductiva del animal, lo que conlleva importantes pérdidas económicas en criaderos de perros y pérdida afectiva para propietarios cuando el animal debe ser eutanasiado. El diagnóstico suele realizarse mediante pruebas serológicas, las cuales presentan como principal desventaja la obtención de resultados falsos positivos por reactividad cruzada con otras bacterias, ya sea del mismo o distinto género, o bien resultados falsos negativos en casos crónicos de infección y por esta razón se requiere de la confirmación diagnóstica mediante el aislamiento bacteriano como método diagnóstico directo. Sin embargo, lo anterior conlleva tanto el riesgo de infección al personal de laboratorio que trabaja con esta bacteria debido a su carácter zoonótico y por otra parte involucra mayor tiempo debido a que este método requiere de un periodo prolongado de incubación. En consideración a lo anterior, esta memoria de título tuvo por objeto desarrollar un PCR convencional capaz de diferenciar entre la detección de B. canis, B. suis y B. abortus, mediante el diseño in silico de un par de partidores que generaron un amplicón de mayor tamaño para la especie de interés en relación a las otras dos especies de Brucella. El análisis de las secuencias nucleotídicas obtenidas mediante los programas de libre acceso Clustal W y BLAST permitió establecer la identidad nucleotídica de los fragmentos obtenidos, corroborando así la detección diferencial entre B. canis, B. suis y B. abortus. Así, este método constituye una prometedora alternativa al aislamiento bacteriano como método diagnóstico directo y complementario a pruebas serológicas.

The parallel gene expression profiles of Brucella melitensis and the host have not been elaborated. In this study, I analyze and discuss the transcriptional profiles of B. melitensis invasive-associated genes, the expression profile of intracellular B. melitensis and B. melitensis-infected non-phagocytic cells in the first 12 h post-infection (PI), and the in vivo temporal global transcriptome of both B. melitensis and the infected bovine host in the first 4 h PI. The initial study found that B. melitensis at late-log phase of growth were more invasive in non-phagocytic cells than at early-log or stationary growth phase. Microarray-based studies identified 454 Brucella genes differentially expressed between the most and the least invasive growth phases. Additionally, B. melitensis strains with transposon interrupted in loci BMEII0380 (acrA) and BMEI1538 (hypothetical protein) were found to be deficient in internalization compare with the wild-type strain. A second experiment was designed with the goal of characterizing host and pathogen transcriptome in parallel. For detecting intracellular Brucella gene expression, a combined protocol consisting of a linear amplification of sense-stranded RNA biased to pathogen transcripts to the previously enriched host:pathogen RNA mixed sample, was developed. RNA samples were hybridized on human and Brucella cDNA microarrays, which analysis revealed a common down-regulation transcriptional profile at 4 h PI that was reverse at 12 h PI. The integrity of B. melitensis virB operon and the expression of host MAPK1 were confirmed as critical for early B. melitensis intracellular survival and replication in non-phagocytic cells. Finally, a temporal morphological and molecular characterization of the initial B. melitensis:bovine host interaction using a calf ileal loop model was performed. B. melitensis was isolated from intestinal Peyer’s patches as soon as 15 min and from systemic blood after 30 min postintra luminal inoculation. Microarray results revealed a common transcriptional profile in Brucella, but two different transcriptional profiles were identified in the host in the first 4 h PI. The importance of differentially expressed biological processes, pathways and individual genes in the initial Brucella pathogenesis is discussed.

Thesis (Ingeniero Zootecnista)--Escuela Superior Politecnica de Chimborazo. Facultad de Ciencias Pecuarias. Escuela de Ingeniera Zootecnica. / Abstract in Spanish and English.

Thesis (M.S.)--University of Wisconsin--Madison, 1949. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 30-33).

Thesis (Ingeniero Zootecnista)--Escuela Superior Politecnica de Chimborazo. Facultad de Ciencias Pecuarias. Escuela de Ingeniera Zootecnica. / Abstract in Spanish and English.

Thesis (M.S.)--Virginia Polytechnic Institute and State University, 1992. / Vita. Abstract. Includes bibliographical references (leaves 33-36). Also available via the Internet.

Thesis (M.S.)--University of Wisconsin--Madison, 1966. / eContent provider-neutral record in process. Description based on print version record. Bibliography: l. 77-84.

München, Univ., Diss., 2005